Spiro compounds and adhesion molecule inhibitors containing the same as the active ingredient

ABSTRACT

Disclosed are novel spiro derivatives and their medical uses, especially as adhesion molecule inhibitors useful for therapies of inflammatory diseases. The spiro derivative according to the present invention has the chemical structure, for example, represented by the following Formula (31):

This application is the national phase under 35 U.S.C. § 371 of PCTInternational Application No. PCT/JP01/08290 which has an Internationalfiling date of Sep. 25, 2001, which designated the United States ofAmerica.

TECHNICAL FIELD

The present invention relates to a novel spiro derivative or apharmaceutically acceptable salt thereof, useful as an adhesion moleculeinhibitor, especially VLA-4 inhibitor, to an adhesion moleculeinhibitor, especially VLA-4 inhibitor, containing the same, and to atherapeutic agent against inflammatory diseases containing the same.

BACKGROUND ART

Adhesion molecules participate in adhesion between cells and cells, andbetween cells and cell matrix. Adhesion molecules include a number offamilies such as integrin family and immunoglobulin super family. Theadhesion molecules belonging to integrin family are those expressed onleukocytes such as lymphocytes, monocytes, basophils and eosinophils.These adhesion molecules have heterodimer structure, in which an a chainand a β chain are non-covalently bound, and are classified into somesubfamilies depending on the species of the β chain. VLA-4 (very lateantigen-4) also called α4β1 or CD49d/CD29, a member of the integrinfamily, participates in the interactions between leukocytes and vascularendothelial cells or extracellular matrix, and participates ininfiltration of leukocytes into inflammatory site. VCAM-1 (vascular celladhesion molecule-1) and fibronectin are known as the adhesion moleculeswhich interact with VLA-4.

The binding site on fibronectin, which binds to VLA-4 is a fibronectinfragment called CS-1. It has been reported that the minimum unitrequired for the binding in this fragment consists of 3 amino acidresidues, that is, Leucine-Aspartic acid-Valine.

Linear or cyclic peptidic VLA-4 adhesion inhibitor compounds based onthe 3 amino acid residues, Leucine-Aspartic acid-Valine have beenreported (WO/15973).

On the other hand, it is known that the expression level of VCAM-1 whichis another adhesion molecule that also interacts with VLA-4, isincreased by stimulation by a cytokine such as IL-1, TNF-α or IL-4, andthat VCAM-1 interacts with VLA-4 existing on cells such as lymphocytes,NK cells, monocytes and eosinophils. VLA-4 and VCAM-1 participate in theinfiltration of leukocytes into inflammatory sites through bloodvessels. From this view point, the interaction between VLA-4 and VCAM-1is very important in inflammatory reaction.

Among the adhesion molecules, VCAM-1 belongs to the immunoglobulin superfamily, and 7-Ig-like-domain VCAM-1 and 6-Ig-like-domain VCAM-1 areknown. Mutation experiments of VCAM-1 revealed that the binding sites onVCAM-1 for binding to VLA-4 are located in domain 1 and domain 4, andthat the amino acid sequence of glutamine-isoleucine-asparticacid-serine-proline on the CD loop is important for the binding to VLA-4(e.g., J. Cell Biol., 124, 601(1994)). J. H. WANG et al. reported acyclic peptide Cys*GlnIleAspSerProCys* (Cys*Cys* represents disulfidebond) which has an inhibitory activity against adhesion of VLA-4, whichcyclic peptide is based on the glutamine-isoleucine-asparticacid-serine-proline (Proc. Natl. Acad. Sci. USA, 92, 5714 (1995)). Lowmolecular compounds having VLA-4-inhibitory activity have also beenreported (e.g., U.S. Pat. No. 5,770,573, U.S. Pat. No. 5,821,231 andWO99/6436).

The fact that VLA-4 plays an important role in inflammatory reaction hasbeen proved by experiments using anti-VLA-4 antibody in animal modelssuch as contact hypersensitivity, delayed type hypersensitivity models(mouse and rat), experimental autoimmune encephalomyelitis models (mouseand rat), nephrotic nephritis (rat), passive cutaneous anaphylaxis model(guinea pig), immunocomplex-induced pulmonary injury model (rat),spontaneous colitis model (monkey), asthma model (sheep) and adjuvantarthritis model.

DISCLOSURE OF THE INVENTION

It has been proved that the cause of development of chronic inflammatorydiseases such as allergic inflammation and chronic rheumatoid arthritisis the repetition of accumulation of leukocytes at the inflammatorysite. However, as the drugs for the therapies of these diseases, drugshaving activities to inhibit actions of chemical mediators, drugs havingactivities to inhibit production of chemical mediators, and drugs havingactivities to inhibit production of active oxygen are conventionallyused. Drugs which inhibit activation of leukocytes, such as steroiddrugs, are also used. Since these drugs do not have an activity toinhibit accumulation of leukocytes to the inflammatory site as theirmain actions, they cannot inhibit development of inflammation. Incontrast, since adhesion molecules VLA-4 and VCAM-1 mainly participatein the process of accumulation of the leukocytes to the inflammatorysite, a novel compound having an activity to inhibit the adhesion ofVLA-4 and VCAM-1 is thought to inhibit the accumulation of theleukocytes to the inflammatory site. Thus, the probability that such acompound is an effective therapeutic drug against the above-mentioneddiseases is high.

An object of the present invention is to discover a compound whichinhibits cell infiltration via adhesion molecules, especially, adhesionmolecule VLA-4, thereby making it possible to prevent and cureinflammatory diseases caused by infiltration of leukocytes such asmonocytes, lymphocytes and eosinophils.

The present inventors intensively studied to discover that specificnovel spiro derivatives and pharmaceutically acceptable salts thereofhave activities to inhibit cell adhesion via adhesion molecules,especially adhesion molecule VLA-4, thereby completing the presentinvention.

That is, the present invention provides a spiro derivative representedby the Formula I:

-   [wherein l and m independently represent integers of 0 to 2; n    represents an integer of 1 to 3; A represents an oxygen atom, carbon    atom or a nitrogen atom (with the proviso that when A is an oxygen    atom, R₃ does not exist); B represents a carbon atom or a nitrogen    atom; C′ and D represent hydrogen, or C′ and D cooperatively    represent carbonyl; X₁ and Y₁ independently represent hydrogen,    halogen, C₁-C₈ alkyl, C₁-C₈ alkoxy, cyano, nitro, hydroxyl, amino or    tetrazole; R₁ and R₂ independently represent hydrogen or C₁-C₆    linear alkyl; R₃ and R₄ independently represent hydrogen, C₁-C₆    linear alkyl, C₃-C₈ branched alkyl, or phenyl or benzyl, this phenyl    or benzyl being substituted with 0 to 2 substituents selected from    the group consisting of halogen, C₁-C₈ alkyl, C₁-C₈ alkoxy, cyano,    nitro, hydroxyl, amino and tetrazole; F represents —CH₂— or —C(O)—;-   when A is a nitrogen atom, R₃, A and R₄ may cooperatively represent-   (i) Formula II:-   (wherein p represents an integer of 0 to 4; E represents a carbon    atom or a nitrogen atom; R₆ and R₇ independently represent hydrogen,    C₁-C₆ linear alkyl, C₃-C₈ branched alkyl, C₁-C₆ linear alkylacyl,    C₃-C₈ branched alkylacyl, pyrrolidinecarbonyl, peperidinecarbonyl,    or phenyl, phenylsulfonyl, benzoyl, benzyl, indole or N-phenylamide,    this phenyl, phenylsulfonyl, benzoyl, benzyl, indole or    N-phenylamide being substituted with 0 to 2 substituents selected    from the group consisting of halogen, C₁-C₈ alkyl, C₁-C₈ alkoxy,    cyano, nitro, hydroxyl, amino and tetrazole, or Formula III:-   (wherein R₈ represents hydrogen, C₁-C₆ linear alkyl, C₃-C₈ branched    alkyl, or phenyl or benzyl, this phenyl or benzyl being substituted    with 0 to 2 substituents selected from the group consisting of    halogen, C₁-C₈ alkyl, C₁-C₈ alkoxy, cyano, nitro, hydroxyl, amino    and tetrazole),-   (ii) Formula IV:-   (wherein R₉ represents C₁-C₆ linear alkyl, C₃-C₈ branched alkyl,    C₁-C₆ linear alkylacyl, C₃-C₈ branched alkylacyl, C₅-C₇    cycloalkylacyl, C₁-C₆ linear alkylsulfonyl, C₃-C₈ branched    alkylsulfonyl, or benzoyl, phenylsulfonyl or benzyl, this benzoyl,    phenylsulfonyl or benzyl being substituted with 0 to 2 substituents    selected from the group consisting of halogen, C₁-C₈ alkyl, C₁-C₈    alkoxy, cyano, nitro, hydroxyl, amino and tetrazole)-   (iii) Formula V:-   (wherein R₁₀ represents hydrogen, C₁-C₆ linear alkyl, C₃-C₈ branched    alkyl, C₆-C₁₀ alkylcycloalkyl, or phenyl or benzyl, this phenyl or    benzyl being substituted with 0 to 2 substituents selected from the    group consisting of halogen, C₁-C₈ alkyl, C₁-C₈ alkoxy, cyano,    nitro, hydroxyl, amino and tetrazole);-   When A is a carbon atom, R₃, A and R₄ may cooperatively form    adamantyl, Formula VI:-   (wherein definitions of X₂ and Y₂ are the same as those of X₁ and    Y₁, respectively) or Formula VII:-   (wherein definition of R₁₁ is the same as that of R₉);-   R₅ represents hydrogen, C₁-C₆ linear alkyl, C₃-C₈ branched alkyl,    allyl, homoallyl, C₆-C₁₀ alkylcycloalkyl, or phenyl, benzyl,    phenethyl, styryl or naphthylmethyl, this phenyl, benzyl, phenethyl,    styryl or naphthylmethyl being substituted with 0 to 2 substituents    selected from the group consisting of halogen, C₁-C₈ alkyl, C₁-C₈    alkoxy, cyano, nitro, hydroxyl, amino and tetrazole]    or a pharmaceutically acceptable salt thereof.

The present invention also provides an adhesion molecule inhibitorcomprising the spiro derivative or a pharmaceutically acceptable saltthereof according to the present invention. The present inventionfurther provides a medical use of the spiro derivative or apharmaceutically acceptable salt thereof according to the presentinvention, and especially, a therapeutic agent for inflammatorydiseases. The present invention still further provides a method forinhibiting an adhesion molecule, comprising administering an effectiveamount of the spiro derivative or a pharmaceutically acceptable saltthereof according to the present invention. The present invention stillfurther provides a use of the spiro derivative or a pharmaceuticallyacceptable salt thereof according to the present invention for theproduction of a pharmaceutical. The present invention still furtherprovides a use of the spiro derivative or a pharmaceutically acceptablesalt thereof according to the present invention for the production of anadhesion molecule inhibitor.

By the present invention, novel substances having activities to inhibitcell adhesion via adhesion molecules, especially adhesion moleculeVLA-4, were provided. By the present invention, prevention and therapyof the inflammatory diseases caused by infiltration of leukocytes suchas monocytes, lymphocytes and eosinophils can be attained.

Best Mode for Carrying Out the Invention

As mentioned above, the spiro derivatives according to the presentinvention are represented by the above-described Formula I. In FormulaI, 1 represents an integer of 0 to 2; m represents an integer of 0 to 2;n represents an integer of 1 to 3; A represents an oxygen atom, carbonatom or a nitrogen atom (with the proviso that when A is an oxygen atom,R₃ does not exist); B represents a carbon atom or a nitrogen atom; C′and D represent hydrogen, or C′ and D cooperatively represent carbonyl;X₁ and Y₁ independently represent hydrogen, halogen, C₁-C₈ alkyl such asmethyl, ethyl, n-propyl or isopropyl, C₁-C₈ alkoxy such as methoxy,ethoxy, n-propyloxy or isopropyloxy, cyano, nitro, hydroxyl, amino ortetrazole; R₁ and R₂ represent hydrogen or C₁-C₆ linear alkyl, that is,methyl, ethyl, n-propyl, n-butyl, n-pentyl or n-hexyl; R₃ and R₄independently represent hydrogen, C₁-C₆ linear alkyl, that is, methyl,ethyl, n-propyl, n-butyl, n-pentyl or n-hexyl, C₃-C₈ branched alkyl suchas 1-methylethyl, 1-methylpropyl, 2-methylpropyl, 1-methylbutyl,2-methylbutyl, 3-methylbutyl, 1-methylpentyl, 2-methylpentyl,3-methylpentyl, 4-methylpentyl, 1-methylhexyl, 2-methylhexyl,3-methylhexyl, 4-methylhexyl, 5-methylhexyl, 3,5-dimethylhexyl,3,6-dimethylhexyl or 4,5-dimethylhexyl, or phenyl or benzyl, this phenylor benzyl being substituted with 0 to 2 substituents selected from thegroup consisting of halogen, C₁-C₈ alkyl, C₁-C₈ alkoxy, cyano, nitro,hydroxyl, amino and tetrazole, such as phenyl, 2-cyanophenyl,2-hydroxyphenyl, 2-chlorophenyl, 2-nitrophenyl, 2-aminophenyl,2-bromophenyl, 2-fluorophenyl, 2-tetrazoylphenyl, 2,6-dihydroxyphenyl,2,6-diemthoxyphenyl, 2,6-dichlorophenyl, 2,6-dinitrophenyl,2,6-dimethylphenyl, benzyl, 2-cyanobenzyl, 2-hydroxybenzyl,2-chlorobenzyl, 2-nitrobenzyl, 2-aminobenzyl, 2-bromobenzyl,2-fluorobenzyl, 2-tetrazoylbenzyl, 2,6-dihydroxybenzyl,2,6-dimethoxybenzyl, 2,6-dichlorobenzyl, 2,6-dinitrobenzyl or2,6-dimethylbenzyl; F represents —CH₂— or —C(O)—;

-   when A is a nitrogen atom, R₃, A and R₄ may cooperatively represent-   (i) Formula II:-   (wherein p represents an integer of 0 to 4, E represents a carbon    atom or a nitrogen atom; R₆ and R₇ independently represent hydrogen,    C₁-C₆ linear alkyl, that is, methyl, ethyl, n-propyl, n-butyl,    n-pentyl or n-hexyl, C₃-C₈ branched alkyl such as 1-methylethyl,    1-methylpropyl, 2-methylpropyl, 1-methylbutyl, 2-methylbutyl,    3-methylbutyl, 1-methylpentyl, 2-methylpentyl, 3-methylpentyl,    4-methylpentyl, 1-methylhexyl, 2-methylhexyl, 3-methylhexyl,    4-methylhexyl, 5-methylhexyl, 3,5-dimethylhexyl, 3,6-dimethylhexyl    or 4,5-dimethylhexyl, C₁-C₆ linear alkylacyl such as acetyl,    ethyloxy or n-propyloxy, C₃-C₈ branched alkylacyl such as    isopropyloxy, isobutyloxy, isopentyloxy or isohexyloxy,    pyrrolidinecarbonyl, peperidinecarbonyl, or phenyl, phenylsulfonyl,    benzoyl, benzyl, indole or N-phenylamide, this phenyl,    phenylsulfonyl, benzoyl, benzyl, indole or N-phenylamide being    substituted with 0 to 2 substituents selected from the group    consisting of halogen, C₁-C₈ alkyl, C₁-C₈ alkoxy, cyano, nitro,    hydroxyl, amino and tetrazole, such as phenyl, 2-cyanophenyl,    2-hydroxyphenyl, 2-chlorophenyl, 2-nitrophenyl, 2-aminophenyl,    2-bromophenyl, 2-fluorophenyl, 2-tetrazoylphenyl,    2,6-dihydroxyphenyl, 2,6-dimethoxyphenyl, 2,6-dichlorophenyl,    2,6-dinitrophenyl, 2,6-dimethylphenyl, benzoyl, 2-cyanobenzoyl,    2-hydroxybenzoyl, 2-chlorobenzoyl, 2-nitrobenzoyl, 2-aminobenzoyl,    2-bromobenzoyl, 2-fluorobenzoyl, 2-tetrazoylbenzoyl,    2,6-dihydroxybenzoyl, 2,6-dimethoxybenzoyl, 2,6-dichlorobenzoyl,    2,6-dinitrobenzoyl, 2,6-dimethylbenzoyl, benzyl, 2-cyanobenzyl,    2-hydroxybenzyl, 2-chlorobenzyl, 2-nitrobenzyl, 2-amionobenzyl,    2-bromobenzyl, 2-fluorobenzyl, 2-tetrazoylbenzyl,    2,6-dihydroxybenzyl, 2,6-dimethoxybenzyl, 2,6-dichlorobenzyl,    2,6-dinitrobenzyl, 2,6-dimethylbenzyl, phenylsulfonyl,    2-cyanophenylsulfonyl, 2-hydroxyphenylsulfonyl,    2-chlorophenylsulfonyl, 2-nitrophenylsulfonyl,    2-aminophenylsulfonyl, 2-bromophenylsulfonyl,    2-fluorophenylsulfonyl, 2-tetrazoylphenylsulfonyl,    2,6-dihydroxyphenylsulfonyl, 2,6-dimethoxyphenylsulfonyl,    2,6-dichlorophenylsulfonyl, 2,6-dinitrophenylsulfonyl,    2,6-dimethylphenylsulfonyl, N-phenylcarboxamide,    N-(2-cyanophenyl)carboxamide, N-(2-cyanophenyl)carboxamide,    N-(2-hydroxyphenyl)carboxamide, N-(2-chlorophenyl)carboxamide,    N-(2-nitrophenyl)carboxamide, N-(2-aminophenyl)carboxamide,    N-(2-bromophenyl)carboxamide, N-(2-fluorophenyl)carboxamide,    N-(2-tetrazoylphenyl)carboxamide,    N-(2,6-dihydroxyphenyl)carboxamide,    N-(2,6-dimethoxyphenyl)carboxamide,    N-(2,6-dichlorophenyl)carboxamide, N-(2,6-dinitrophenyl)carboxamide    or N-(2,6-dimethylphenyl)carboxamide, or Formula III:-   (wherein R₈ represents hydrogen, C₁-C₆ linear alkyl, that is,    methyl, ethyl, n-propyl, n-butyl, n-pentyl or n-hexyl, C₃-C₈    branched alkyl such as 1-methylethyl, 1-methylpropyl,    2-methylpropyl, 1-methylbutyl, 2-methylbutyl, 3-methylbutyl,    1-methylpentyl, 2-methylpentyl, 3-methylpentyl, 4-methylpentyl,    1-methylhexyl, 2-methylhexyl, 3-methylhexyl, 4-methylhexyl,    5-methylhexyl, 3,5-dimethylhexyl, 3,6-dimethylhexyl or    4,5-dimethylhexyl, or phenyl or benzyl, this phenyl or benzyl being    substituted with 0 to 2 substituents selected from the group    consisting of halogen, C₁-C₈ alkyl, C₁-C₈ alkoxy, cyano, nitro,    hydroxyl, amino and tetrazole, such as phenyl, 2-cyanophenyl,    2-hydroxyphenyl, 2-chlorophenyl, 2-nitrophenyl, 2-aminophenyl,    2-bromophenyl, 2-fluorophenyl, 2-tetrazoylphenyl,    2,6-dihydroxyphenyl, 2,6-dimethoxyphenyl, 2,6-dichlorophenyl,    2,6-dinitrophenyl, 2,6-dimethylphenyl, benzyl, 2-cyanobenzyl,    2-hydroxybenzyl, 2-chlorobenzyl, 2-nitrobenzyl, 2-aminobenzyl,    2-bromobenzyl, 2-fluorobenzyl, 2-tetrazoylbenzyl,    2,6-dihydroxybenzyl, 2,6-dimethoxybenzyl, 2,6-dichlorobenzyl,    2,6-dinitrobenzyl or 2,6-dimethylbenzyl),-   (ii) Formula IV:-   (wherein R₉ represents hydrogen, C₁-C₆ linear alkyl, that is,    methyl, ethyl, n-propyl, n-butyl, n-pentyl or n-hexyl, C₃-C₈    branched alkyl such as 1-methylethyl, 1-methylpropyl,    2-methylpropyl, 1-methylbutyl, 2-methylbutyl, 3-methylbutyl,    1-methylpentyl, 2-methylpentyl, 3-methylpentyl, 4-methylpentyl,    1-methylhexyl, 2-methylhexyl, 3-methylhexyl, 4-methylhexyl,    5-methylhexyl, 3,5-dimethylhexyl, 3,6-dimethylhexyl or    4,5-dimethylhexyl, C₁-C₆ linear alkylacyl, that is, acetyl,    propionyl, butyryl, valeryl, pentanoyl or hexanoyl, C₃-C₈ branched    alkylacyl such as isopropionyl, isobutyryl, pivaloyl, isopentanoyl,    isohexanoyl or isoheptanoyl, C₅-C₇ cycloalkylacyl such as    cyclopentylcarbonyl or cyclohexylcarbonyl, C₁-C₆ linear    alkylsulfonyl such as mesyl, ethanesulfonyl, n-propansulfonyl,    n-butanesulfonyl, n-pentanesulfonyl or n-hexanesulfonyl, C₃-C₈    branched alkylsulfonyl such as isopropanesulfonyl,    isobutanesulfonyl, t-butanesulfonyl, isopentanesulfonyl,    isohexanesulfonyl or isoheptanesulfonyl; or benzoyl, phenylsulfonyl    or benzyl, this benzoyl, phenylsulfonyl or benzyl being substituted    with 0 to 2 substituents selected from the group consisting of    halogen, C₁-C₈ alkyl, C₁-C₈ alkoxy, cyano, nitro, hydroxyl, amino    and tetrazole, such as benzoyl, 2-cyanobenzoyl, 2-hydroxybenzoyl,    2-chlorobenzoyl, 2-nitrobenzoyl, 2-aminobenzoyl, 2-bromobenzoyl,    2-fluorobenzoyl, 2-tetrazoylbenzoyl, 2,6-dihydroxybenzoyl,    2,6-dimethoxybenzoyl, 2,6-dichlorobenzoyl, 2,6-dinitrobenzoyl,    2,6-dimethylbenzoyl, phenylsulfonyl, 2-cyanophenylsulfonyl,    2-hydroxyphenylsulfonyl, 2-chlorophenylsulfonyl,    2-nitrophenylsulfonyl, 2-aminophenylsulfonyl, 2-bromophenylsulfonyl,    2-fluorophenylsulfonyl, 2-tetrazoylphenylsulfonyl,    2,6-dihydroxyphenylsulfonyl, 2,6-dimethoxyphenylsulfonyl,    2,6-dichlorophenylsulfonyl, 2,6-dinitrophenylsulfonyl,    2,6-dimethylphenylsulfonyl, benzyl, 2-cyanobenzyl, 2-hydroxybenzyl,    2-chlorobenzyl, 2-nitrobenzyl, 2-aminobenzyl, 2-bromobenzyl,    2-fluorobenzyl, 2-tetrazoylbenzyl, 2,6-dihydroxybenzyl,    2,6-dimethoxybenzyl, 2,6-dichlorobenzyl, 2,6-dinitrobenzyl or    2,6-dimethylbenzyl)-   (iii) Formula V:-   (wherein R₁₀ represents hydrogen, C₁-C₆ linear alkyl, that is,    methyl, ethyl, n-propyl, n-butyl, n-pentyl or n-hexyl, C₃-C₈    branched alkyl such as 1-methylethyl, 1-methylpropyl,    2-methylpropyl, 1-methylbutyl, 2-methylbutyl, 3-methylbutyl,    1-methylpentyl, 2-methylpentyl, 3-methylpentyl, 4-methylpentyl,    1-methylhexyl, 2-methylhexyl, 3-methylhexyl, 4-methylhexyl,    5-methylhexyl, 3,5-dimethylhexyl, 3,6-dimethylhexyl or    4,5-dimethylhexyl, C₆-C₁₀ alkylcycloalkyl such as cyclopentylmethyl,    cyclohexylmethyl or cycloheptylmethyl, or phenyl or benzyl, this    phenyl or benzyl being substituted with 0 to 2 substituents selected    from the group consisting of halogen, C₁-C₈ alkyl, C₁-C₈ alkoxy,    cyano, nitro, hydroxyl, amino and tetrazole, such as phenyl,    2-cyanophenyl, 2-hydroxyphenyl, 2-chlorophenyl, 2-nitrophenyl,    2-aminophenyl, 2-bromophenyl, 2-fluorophenyl, 2-tetrazoylphenyl,    2,6-dihydroxyphenyl, 2,6-dimethoxyphenyl, 2,6-dichlorophenyl,    2,6-dinitrophenyl, 2,6-dimethylphenyl, benzyl, 2-cyanobenzyl,    2-hydroxybenzyl, 2-chlorobenzyl, 2-nitrobenzyl, 2-aminobenzyl,    2-bromobenzyl, 2-fluorobenzyl, 2-tetrazoylbenzyl,    2,6-dihydroxybenzyl, 2,6-dimethoxybenzyl, 2,6-dichlorobenzyl,    2,6-dinitrobenzyl or 2,6-dimethylbenzyl);-   When A is a carbon atom, R₃, A and R₄ may cooperatively form    adamantyl, Formula VI:-   (wherein definitions of X₂ and Y₂ are the same as those of X₁ and    Y₁, respectively) or Formula VII:-   (wherein definition of R₁₁ is the same as that of R₉);-   R₅ represents hydrogen, C₁-C₆ linear alkyl, that is, methyl, ethyl,    n-propyl, n-butyl, n-pentyl or n-hexyl, C₃-C₈ branched alkyl such as    1-methylethyl, 1-methylpropyl, 2-methylpropyl, 1-methylbutyl,    2-methylbutyl, 3-methylbutyl, 1-methylpentyl, 2-methylpentyl,    3-methylpentyl, 4-methylpentyl, 1-methylhexyl, 2-methylhexyl,    3-methylhexyl, 4-methylhexyl, 5-methylhexyl, 3,5-dimethylhexyl,    3,6-dimethylhexyl or 4,5-dimethylhexyl, allyl, homoallyl, C₆-C₁₀    alkylcycloalkyl such as cyclopentylmethyl, cyclohexylmethyl or    cycloheptylmethyl, or phenyl, benzyl, phenethyl, styryl or naphthyl,    this phenyl, benzyl, phenethyl, styryl or naphthyl being substituted    with 0 to 2 substituents selected from the group consisting of    halogen, C₁-C₈ alkyl, C₁-C₈ alkoxy, cyano, nitro, hydroxyl, amino    and tetrazole, such as phenyl, 2-cyanophenyl, 2-hydroxyphenyl,    2-chlorophenyl, 2-nitrophenyl, 2-aminophenyl, 2-bromophenyl,    2-fluorophenyl, 2-tetrazoylphenyl, 2,6-dihydroxyphenyl,    2,6-dimethoxyphenyl, 2,6-dichlorophenyl, 2,6-dinitrophenyl,    2,6-dimethylphenyl, benzyl, 2-cyanobenzyl, 2-hydroxybenzyl,    2-chlorobenzyl, 2-nitrobenzyl, 2-aminobenzyl, 2-bromobenzyl,    2-fluorobenzyl, 2-tetrazoylbenzyl, 2,6-dihydroxybenzyl,    2,6-dimethoxybenzyl, 2,6-dichlorobenzyl, 2,6-dinitrobenzyl,    2,6-dimethylbenzyl, phenethyl, 2-cyanophenethyl, 2-hydroxyphenethyl,    2-chlorophenethyl, 2-nitrophenethyl, 2-aminophenethyl,    2-bromophenethyl, 2-fluorophenethyl, 2-tetrazoylphenethyl,    2,6-dihydroxyphenethyl, 2,6-dimethoxyphenethyl,    2,6-dichlorophenethyl, 2,6-dinitrophenethyl, 2,6-dimethylphenethyl,    styryl, 2-cyanostyryl, 2-hydroxystyryl, 2-chlorostyryl,    2-nitrostyryl, 2-aminostyryl, 2-bromostyryl, 2-fluorostyryl,    2-tetrazoylstyryl, 2,6-dihydroxystyryl, 2,6-dimethoxystyryl,    2,6-dichlorostyryl, 2,6-dinitrostyryl, 2,6-dimethylstyryl, naphthyl,    2-cyanonaphthyl, 2-hydroxynaphthyl, 2-chloronaphthyl,    2-nitronaphthyl, 2-aminonaphthyl, 2-bromonaphthyl, 2-fluoronaphthyl,    2-tetrazoylnaphthyl, 2,8-dihydroxynaphthyl, 2,8-dimethoxynaphthyl,    2,8-dichloronaphthyl, 2,8-dinitronaphthyl or 2,8-dimethylnaphthyl.

Among the compounds represented by said Formula I, those wherein Frepresents —C(O)—; A and B independently represent carbon atom ornitrogen atom; C′ and D represent hydrogen or C′ and D cooperativelyrepresent carbonyl; X and Y independently represent hydrogen, halogen,methyl, methoxy, cyano, nitro, hydroxyl, amino or tetrazole; R₁ and R₂independently represent hydrogen or C₁-C₆ linear alkyl; R₃ and R₄independently represent hydrogen, C₁-C₆ linear alkyl or C₃-C₈ branchedalkyl, or phenyl or benzyl, this phenyl or benzyl being substituted with0 to 2 substituents selected from the group consisting of halogen,methyl, methoxy, cyano, nitro, hydroxyl, amino and tetrazole;

-   when A is a nitrogen atom, R₃, A and R₄ may cooperatively represent-   (i) said Formula II (wherein p represents an integer of 0 to 4; E    represents a carbon atom or a nitrogen atom; R₆ and R₇ independently    represent hydrogen, C₁-C₆ linear alkyl, C₃-C₈ branched alkyl, C₁-C₆    linear alkylacyl, C₃-C₈ branched alkylacyl, pyrrolidinecarbonyl,    piperidinecarbonyl, or phenyl, phenylsulfonyl, benzoyl, benzyl,    indole or N-phenylamide, this phenyl, phenylsulfonyl, benzoyl,    benzyl, indole or N-phenylamide being substituted with 0 to 2    substituents selected from the group consisting of halogen, methyl,    methoxy, cyano, nitro, hydroxyl, amino and tetrazole, or said    Formula III (wherein R₈ represents hydrogen, C₁-C₆ linear alkyl,    C₃-C₈ branched alkyl, or phenyl or benzyl, this phenyl or benzyl    being substituted with 0 to 2 substituents selected from the group    consisting of halogen, methyl, methoxy, cyano, nitro, hydroxyl,    amino and tetrazole),-   (ii) said Formula IV (wherein R₉ represents C₁-C₆ linear alkyl,    C₃-C₈ branched alkyl, C₁-C₆ linear alkylacyl, C₃-C₈ branched    alkylacyl, C₅-C₇ cycloalkylacyl, C₁-C₆ linear alkylsulfonyl, C₃-C₈    branched alkylsulfonyl, or benzoyl, phenylsulfonyl or benzyl, this    benzoyl, phenylsulfonyl or benzyl being substituted with 0 to 2    substituents selected from the group consisting of halogen, methyl,    methoxy, cyano, nitro, hydroxyl, amino and tetrazole),-   (iii) said Formula V:-   (wherein R₁₀ represents hydrogen, C₁-C₆ linear alkyl, C₃-C₈ branched    alkyl, C₆-C₁₀ alkylcycloalkyl, or phenyl or benzyl, this phenyl or    benzyl being substituted with 0 to 2 substituents selected from the    group consisting of halogen, methyl, methoxy, cyano, nitro,    hydroxyl, amino and tetrazole);-   when A is a carbon atom, R₃, A and R₄ may cooperatively represent    adamantyl or said Formula VI (wherein X₂ and Y₂ represent the same    definitions as described above); R₅ represents hydrogen, C₁-C₆    linear alkyl, C₃-C₈ branched alkyl, allyl, homoallyl, C₆-C₁₀    alkylcycloalkyl, or phenyl, benzyl, phenethyl, styryl or    naphthylmethyl, this phenyl, benzyl, phenethyl, styryl or    naphthylmethyl being substituted with 0 to 2 substituents selected    from the group consisting of halogen, methyl, methoxy, cyano, nitro,    hydroxyl, amino and tetrazole, are also preferred.

Specific examples of the compounds according to the present inventioninclude

-   3-((2,4,8-triaza-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)-2-((2,6-dimethoxyphenyl)carbonylamino)propanoic    acid,-   3-((2,4,8-triaza-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)-2-((2,6-dimethylphenyl)carbonylamino)propanoic    acid,-   3-((2,4,8-triaza-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)-2-((2,6-dichlorophenyl)carbonylamino)propanoic    acid,-   3-((2,4,8-triaza-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)-2-((2,6-difulorophenyl)carbonylamino)propanoic    acid,-   3-((2,4,8-triaza-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)-2-((2-bromo-6-methylphenyl)carbonylamino)propanoic    acid,-   3-((2,4,8-triaza-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)-2-((2-methyl-5-nitrophenyl)carbonylamino)propanoic    acid,-   2-((2,6-dichlorophenyl)carbonylamino)-3-((2,4,8-triaza-2-methyl-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)propanoic    acid,-   2-((2,6-dimethylphenyl)carbonylamino)-3-((2,4,8-triaza-2-methyl-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)propanoic    acid,-   2-((2,6-dimethoxyphenyl)carbonylamino)-3-((2,4,8-triaza-2-methyl-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)propanoic    acid,-   2-((2,6-difluorophenyl)carbonylamino)-3-((2,4,8-triaza-2-methyl-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)propanoic    acid,-   2-((2-bromo-6-methylphenyl)carbonylamino)-3-((2,4,8-triaza-2-methyl-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)propanoic    acid,-   2-((2-methyl-5-nitrophenyl)carbonylamino)-3-((2,4,8-triaza-2-methyl-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)propanoic    acid,-   2-((2,6-dichlorophenyl)carbonylamino)-3-((2,4,8-triaza-2-ethyl-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)propanoic    acid,-   2-((2,6-dimethylphenyl)carbonylamino)-3-((2,4,8-triaza-2-ethyl-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)propanoic    acid,-   2-((2,6-dimethoxyphenyl)carbonylamino)-3-((2,4,8-triaza-2-ethyl-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)propanoic    acid,-   2-((2,6-difluorophenyl)carbonylamino)-3-((2,4,8-triaza-2-ethyl-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)propanoic    acid,-   2-((2-bromo-6-methylphenyl)carbonylamino)-3-((2,4,8-triaza-2-ethyl-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)propanoic    acid,-   2-((2-methyl-5-nitrophenyl)carbonylamino)-3-((2,4,8-triaza-2-ethyl-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)propanoic    acid,-   2-((2,6-dichlorophenyl)carbonylamino)-3-((2,4,8-triaza-1-oxo-4-phenyl-2-propylspiro[4.5]dec-8-yl)carbonylamino)propanoic    acid,-   2-((2,6-dimethylphenyl)carbonylamino)-3-((2,4,8-triaza-1-oxo-4-phenyl-2-propylspiro[4.5]dec-8-yl)carbonylamino)propanoic    acid,-   2-((2,6-dimethoxyphenyl)carbonylamino)-3-((2,4,8-triaza-1-oxo-4-phenyl-2-propylspiro[4.5]dec-8-yl)carbonylamino)propanoic    acid,-   2-((2,6-difluorophenyl)carbonylamino)-3-((2,4,8-triaza-1-oxo-4-phenyl-2-propylspiro[4.5]dec-8-yl)carbonylamino)propanoic    acid,-   2-((2-bromo-6-methylphenyl)carbonylamino)-3-((2,4,8-triaza-1-oxo-4-phenyl-2-propylspiro[4.5]dec-8-yl)carbonylamino)propanoic    acid,-   2-((2-methyl-5-nitrophenyl)carbonylamino)-3-((2,4,8-triaza-1-oxo-4-phenyl-2-propylspiro[4.5]dec-8-yl)carbonylamino)propanoic    acid,-   2-((2,6-dimethoxyphenyl)carbonylamino)-3-((2,4,8-triaza-2-(methylethyl)-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)propanoic    acid,-   2-((2,6-dimethylphenyl)carbonylamino)-3-((2,4,8-triaza-2-(methylethyl)-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)propanoic    acid,-   2-((2,6-dichlorophenyl)carbonylamino)-3-((2,4,8-triaza-2-(methylethyl)-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)propanoic    acid,-   2-((2,6-difluorophenyl)carbonylamino)-3-((2,4,8-triaza-2-(methylethyl)-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)propanoic    acid,-   2-((2-bromo-6-methylphenyl)carbonylamino)-3-((2,4,8-triaza-2-(methylethyl)-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)propanoic    acid,-   2-((2-methyl-5-nitrophenyl)carbonylamino)-3-((2,4,8-triaza-2-(methylethyl)-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)propanoic    acid,-   2-((2,6-dimethoxyphenyl)carbonylamino)-3-((2,4,8-triaza-2-(2-methylpropyl)-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)propanoic    acid,-   2-((2,6-dimethylphenyl)carbonylamino)-3-((2,4,8-triaza-2-(2-methylpropyl)-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)propanoic    acid,-   2-((2,6-dichlorophenyl)carbonylamino)-3-((2,4,8-triaza-2-(2-methylpropyl)-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)propanoic    acid,-   2-((2-methyl-5-nitrophenyl)carbonylamino)-3-((2,4,8-triaza-2-(2-methylpropyl)-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)propanoic    acid,-   2-((2-bromo-6-methylphenyl)carbonylamino)-3-((2,4,8-triaza-2-(2-methylpropyl)-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)propanoic    acid,-   2-((2,6-difluorophenyl)carbonylamino)-3-((2,4,8-triaza-2-(2-methylpropyl)-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)propanoic    acid,-   2-((2,6-dimethoxyphenyl)carbonylamino)-3-((2,4,8-triaza-1-oxo-4-phenyl-2-benzylspiro[4.5]dec-8-yl)carbonylamino)propanoic    acid,-   2-((2,6-dimethylphenyl)carbonylamino)-3-((2,4,8-triaza-1-oxo-4-phenyl-2-benzylspiro[4.5]dec-8-yl)carbonylamino)propanoic    acid,-   2-((2,6-dichlorophenyl)carbonylamino)-3-((2,4,8-triaza-1-oxo-4-phenyl-2-benzylspiro[4.5]dec-8-yl)carbonylamino)propanoic    acid,-   2-((2,6-difluorophenyl)carbonylamino)-3-((2,4,8-triaza-1-oxo-4-phenyl-2-benzylspiro[4.5]dec-8-yl)carbonylamino)propanoic    acid,-   2-((2-bromo-6-methylphenyl)carbonylamino)-3-((2,4,8-triaza-1-oxo-4-phenyl-2-benzylspiro[4.5]dec-8-yl)carbonylamino)propanoic    acid,-   2-((2-methyl-5-nitrophenyl)carbonylamino)-3-((2,4,8-triaza-1-oxo-4-phenyl-2-benzylspiro[4.5]dec-8-yl)carbonylamino)propanoic    acid,-   2-((2,6-dimethoxyphenyl)carbonylamino)-3-((2,4,8-triaza-2-((2,6-dimethylphenyl)methyl)-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)propanoic    acid,-   2-((2,6-dimethylphenyl)carbonylamino)-3-((2,4,8-triaza-2-((2,6-dimethylphenyl)methyl)-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)propanoic    acid,-   2-((2,6-dichlorophenyl)carbonylamino)-3-((2,4,8-triaza-2-((2,6-dimethylphenyl)methyl)-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)propanoic    acid,-   2,3-di((2,4,8-triaza-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)propanoic    acid,-   2,3-di((2,4,8-triaza-2-methyl-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)propanoic    acid,-   2,3-di((2,4,8-triaza-1-oxo-4-phenyl-2-benzylspiro[4.5]dec-8-yl)carbonylamino)propanoic    acid,-   2,3-di((2,4,8-triaza-1-oxo-4-phenyl-2-propylspiro[4.5]dec-8-yl)carbonylamino)propanoic    acid,-   2,3-di((2,4,8-triaza-2-(2-methylpropyl)-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)propanoic    acid,-   2,3-di((2,4,8-triaza-2-(2-methylethyl)-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)propanoic    acid,-   3-((2,4,8-triaza-2-methyl-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)-2-((2,4,8-triaza-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)propanoic    acid,-   3-((2,4,8-triaza-2-(2-methylpropyl)-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)-2-((2,4,8-triaza-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)propanoic    acid,-   3-((2,4,8-triaza-2-(2-methylethyl)-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)-2-((2,4,8-triaza-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)propanoic    acid,-   3-((2,4,8-triaza-1-oxo-4-phenyl-2-benzylspiro[4.5]dec-8-yl)carbonylamino)-2-((2,4,8-triaza-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)propanoic    acid,-   2-((4-(2-methylpropanoylamino)-4-phenylpiperidyl)carbonylamino)-3-((2,4,8-triaza-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)propanoic    acid,-   2-((4-(2-methylpropanoylamino)-4-phenylpiperidyl)carbonylamino)-3-((2,4,8-triaza-2-(2-methylpropyl)-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)propanoic    acid,-   2-((4-(2-methylpropanoylamino)-4-phenylpiperidyl)carbonylamino)-3-((2,4,8-triaza-2-methyl-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)propanoic    acid,-   2-((4-(2-methylpropanoylamino)-4-phenylpiperidyl)carbonylamino)-3-((2,4,8-triaza-2-ethyl-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)propanoic    acid,-   2-((4-(2-methylpropanoylamino)-4-phenylpiperidyl)carbonylamino)-3-((2,4,8-triaza-1-oxo-4-phenyl-2-propylspiro[4.5]dec-8-yl)carbonylamino)propanoic    acid,-   2-((4-(2-methylpropanoylamino)-4-phenylpiperidyl)carbonylamino)-3-((2,4,8-triaza-1-oxo-4-phenyl-2-benzylspiro[4.5]dec-8-yl)carbonylamino)propanoic    acid,-   2-((4-(acetylamino)-4-phenylpiperidyl)carbonylamino)-3-((2,4,8-triaza-2-(2-methylpropyl)-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)propanoic    acid,-   2-((4-(acetylamino)-4-phenylpiperidyl)carbonylamino)-3-((2,4,8-triaza-2-methyl-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)propanoic    acid,-   2-((4-(acetylamino)-4-phenylpiperidyl)carbonylamino)-3-((2,4,8-triaza-2-ethyl-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)propanoic    acid,-   2-((4-(acetylamino)-4-phenylpiperidyl)carbonylamino)-3-((2,4,8-triaza-1-oxo-4-phenyl-2-propylspiro[4.5]dec-8-yl)carbonylamino)propanoic    acid,-   2-((4-(acetylamino)-4-phenylpiperidyl)carbonylamino)-3-((2,4,8-triaza-1-oxo-4-phenyl-2-benzylspiro[4.5]dec-8-yl)carbonylamino)propanoic    acid,-   2-((4-phenyl-4-(propanoylamino)piperidyl)carbonylamino)-3-((2,4,8-triaza-2-(2-methylpropyl)-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)propanoic    acid,-   2-((4-phenyl-4-(propanoylamino)piperidyl)carbonylamino)-3-((2,4,8-triaza-2-methyl-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)propanoic    acid,-   2-((4-phenyl-4-(propanoylamino)piperidyl)carbonylamino)-3-((2,4,8-triaza-2-ethyl-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)propanoic    acid,-   2-((4-phenyl-4-(propanoylamino)piperidyl)carbonylamino)-3-((2,4,8-triaza-1-oxo-4-phenyl-2-propylspiro[4.5]dec-8-yl)carbonylamino)propanoic    acid,-   2-((4-phenyl-4-(propanoylamino)piperidyl)carbonylamino)-3-((2,4,8-triaza-1-oxo-4-phenyl-2-benzylspiro[4.5]dec-8-yl)carbonylamino)propanoic    acid,-   2-((4-(2-oxo(3-hydrobenzimidazolyl))piperidyl)carbonylamino)-3-(2,4,8-triaza-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)propanoic    acid,-   2-((4-(2-oxo(3-hydrobenzimidazolyl))piperidyl)carbonylamino)-3-(2,4,8-triaza-1-oxo-4-phenyl-2-benzylspiro[4.5]dec-8-yl)carbonylamino)propanoic    acid,-   2-((4-(2-oxo(3-hydrobenzimidazolyl))piperidyl)carbonylamino)-3-(2,4,8-triaza-1-oxo-4-phenyl-2-propylspiro[4.5]dec-8-yl)carbonylamino)propanoic    acid,-   2-((4-(2-oxo(3-hydrobenzimidazolyl))piperidyl)carbonylamino)-3-(2,4,8-triaza-2-methyl-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)propanoic    acid,-   2-((4-(2-oxo(3-hydrobenzimidazolyl))piperidyl)carbonylamino)-3-(2,4,8-triaza-2-ethyl-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)propanoic    acid,-   2-((4-(2-oxo(3-hydrobenzimidazolyl))piperidyl)carbonylamino)-3-(2,4,8-triaza-2-(2-methylpropyl)-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)propanoic    acid,-   2-(adamantan-2-ylcarbonylamino)-3-((2,4,8-triaza-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)propanoic    acid,-   2-(adamantan-2-ylcarbonylamino)-3-((2,4,8-triaza-1-oxo-4-phenyl-2-benzylspiro[4.5]dec-8-yl)carbonylamino)propanoic    acid,-   2-(adamantan-2-ylcarbonylamino)-3-((2,4,8-triaza-1-oxo-4-phenyl-2-propylspiro[4.5]dec-8-yl)carbonylamino)propanoic    acid,-   2-(adamantan-2-ylcarbonylamino)-3-((2,4,8-triaza-2-methyl-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)propanoic    acid,-   2-(adamantan-2-ylcarbonylamino)-3-((2,4,8-triaza-2-ethyl-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)propanoic    acid,-   2-(adamantan-2-ylcarbonylamino)-3-((2,4,8-triaza-2-(2-methylpropyl)-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)propanoic    acid,-   2-((2,6-dichlorophenyl)amino)carbonylamino)-3-((2,4,8-triaza-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)propanoic    acid,-   2-((2,6-dimethylphenyl)amino)carbonylamino)-3-((2,4,8-triaza-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)propanoic    acid,-   2-((2,6-dimethoxyphenyl)amino)carbonylamino)-3-((2,4,8-triaza-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)propanoic    acid,-   2-((2,6-dichlorophenyl)amino)carbonylamino)-3-((2,4,8-triaza-1-oxo-4-phenyl-2-benzylspiro[4.5]dec-8-yl)carbonylamino)propanoic    acid,-   2-((2,6-dimethylphenyl)amino)carbonylamino)-3-((2,4,8-triaza-1-oxo-4-phenyl-2-benzylspiro[4.5]dec-8-yl)carbonylamino)propanoic    acid,-   2-((2,6-dimethoxyphenyl)amino)carbonylamino)-3-((2,4,8-triaza-1-oxo-4-phenyl-2-benzylspiro[4.5]dec-8-yl)carbonylamino)propanoic    acid,-   2-((2,6-dichlorophenyl)amino)carbonylamino)-3-((2,4,8-triaza-2-(2-methylpropyl)-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)propanoic    acid,-   2-((2,6-dimethylphenyl)amino)carbonylamino)-3-((2,4,8-triaza-2-(2-methylpropyl)-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)propanoic    acid, and-   2-((2,6-dimethoxyphenyl)amino)carbonylamino)-3-((2,4,8-triaza-2-(2-methylpropyl)-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)propanoic    acid,

The processes for producing the compounds represented by Formula I(hereinafter, for example, “the compounds represented by Formula I” mayalso be indicated simply as “Formula I”) will now be described. However,the process for producing each of the compounds is not restricted tothat described herein. In the various production processes, the reactionconditions may be appropriately selected from those described below.

Among the compounds represented by Formula I, those wherein l=0, m=1,n=1, A is a carbon atom, B is a nitrogen atom, C′ and D cooperativelyform carbonyl, F is —C(O)—, R₁ and R₂ are hydrogen atoms, and R₃, A andR₄ cooperatively form Formula VI, that is, those represented by FormulaVIII, or cooperatively form Formula VII, that is, those represented byFormula IX, or A is an oxygen atom, that is, those represented byFormula X:

-   (wherein X₁, X₂, Y₁, Y₂, R₄, R₅ and R₁₁ represent the same meanings    as described above) may be produced by hydrolyzing Formula XI, XII    or XIII:-   (wherein X₁, X₂, Y₁, Y₂, R₄, R₅ and R₁₁ represent the same meanings    as described above) by a base such as aqueous sodium hydroxide    solution or aqueous barium hydroxide solution in an alcoholic    solvent such as methanol. The hydrolysis by the base such as aqueous    sodium hydroxide solution or aqueous barium hydroxide solution may    usually be carried out at about 0° C. to room temperature for about    1 hour to 24 hours, although the reaction conditions are not    restricted thereto. The amount of the base to be added is not    restricted, and may usually be about 1 to 4 equivalents with respect    to Formula XI, XII or XIII.

Formula XI, XII and XIII may be produced from Formula XIV, XV and XVI:

-   (wherein X₂, Y₂, R₄, and R₁₁ represent the same meanings as    described above, Z₁ represents chloro, bromo or hydroxyl),    respectively, and Formula XVII:-   (wherein X₁, Y₁ and R₅ represent the same meanings as described    above).

In cases where one represented by Formula XIV wherein Z₁ is chloro orbromo, or one represented by Formula XVI is used, the desired productmay be produced by reacting Formula XIV or Formula XVI with Formula XVIIin a solvent such as tetrahydrofuran, dimethylformamide, chloroform,dichloromethane or 1,4-dioxane in the presence of a tertiary amine suchas triethylamine or diisopropylamine. The reaction between Formula XIVor XVI and XVII may usually be carried out at about 0° C. to roomtemperature for about 1 hour to 24 hours, although the reactionconditions are not restricted thereto. The mixing ratio (molar ratio;unless otherwise specified, the term “mixing ratio” means molar ratio inthe description below) of Formula XIV or XVI to Formula XVII may be,although not restricted, about 1:1 to 2:1, and the amount of thetertiary amine to be added is, although not restricted, about 1 to 4equivalents with respect to Formula XIV or XVI.

In cases where a compound of Formula XIV wherein Z₁ is hydroxyl, orFormula XV is used, usually, a condensing agent such asdicyclohexylcarbodiimide (DCC),benzotriazol-1-yloxytris(dicyclopentylamino)phosphoniumhexafluorophosphite salt (PyBOP),benzotriazol-1-yloxytris(dimethylamino)phosphoniumhexafluoro phosphitesalt (BOP), diphenylphosphoryl azide (DPPA) or1-ethyl-3-[3-(dimethylamino))propyl]carbodiimide (WSC) is used in asolvent such as tetrahydrofuran, dimethylformamide, chloroform ordichloromethane in the presence of a tertiary amine such astriethylamine, diisopropylamine or N-methylmorpholine. The amount ofsuch a condensing agent to be added is not restricted, and usually about1 to 3 equivalents with respect to Formula XIV or XV. Addition of anadditive such as 1-hydroxybenzotriazole (HOBT) may be advantageous inthe proceeding of the reaction in some cases.

Formula XI, XII and XIII may also be produced by treating Formula XVIII

-   (wherein X₁, Y₁, and R₅ represent the same meanings as described    above)-   with usually 0.5 to 2 equivalents of diphosgene, triphosgene,    1,1-carbonyldiimidazole or the like in a solvent such as    dichloromethane or chloroform, in the presence of a tertiary amine    such as triethylamine or diisopropylamine, and by reacting the    obtained product with Formula XX, XXI and XXII, respectively. The    mixing ratio of Formula XVIII to Formula XX, XXI or XXII may usually    be, although not restricted, about 1:1 to 2:1, and the reaction may    usually be carried out at about 0° C. to room temperature for about    1 hour to 24 hours. The amount of the tertiary amine is not    restricted, and is usually about 1 to 4 equivalents with respect to    diphosgene, triphosgene or 1,1-carbonyldiimidazole. Alternatively,    the desired product may be obtained by reacting Formula XX, XXI or    XXII with Formula XIX in a solvent such as dimethylformamide,    tetrahydrofuran or dimethoxyethane at about 0° C. to room    temperature for about 1 to 24 hours. The mixing ratio of Formula XX,    XXI or XXII to XIX is not restricted, and usually about 1:1 to 1:3.    Alternatively, the desired product may be obtained by reacting    Formula XX, XXI or XXII with p-nitrophenyl chloroformate or phenyl    chloroformate in a solvent such as acetonitrile, dichloromethane or    dimethoxyethane, in the presence of a base such as sodium hydrogen    carbonate or tribenzylamine (first step); and then reacting the    obtained product with Formula XVIII in a solvent such as    acetonitrile, dichloromethane or dimethoxyethane, in the presence of    a tertiary amine such as triethylamine or diisopropylamine (second    step). In this method, the amount of the base such as sodium    hydrogen carbonate or tribenzylamine used in the reaction is not    restricted, and usually about 1 to 4 equivalents with respect to    p-nitrophenyl chloroformate or phenyl chloroformate. The amount of    the tertiary amine to be added is not restricted, and usually about    1 to 4 equivalents with respect to Formula XVIII.

In the first step, the reaction temperature of the reaction betweenFormula XX, XXI or XXII and p-nitrophenyl chloroformate or phenylchloroformate is not restricted, and usually about 0° C. to roomtemperature. The reaction temperature in the second step may usually beabout 0° C. to 50° C. when p-nitrophenyl chloroformate is used, and mayusually be about room temperature to refluxing temperature when phenylchloroformate is used.

Formula XVII may be produced by the following steps:

-   (wherein X₁, Y₁ and R₅ represent the same meanings as described    above).

Step 1 may be carried out in the same manner as in the reaction betweenFormula XVIII or XIX and XX, XXI or XXII.

Step 2 is the step of removing t-butoxycarbonyl group (referred to as“Boc” for short) on the nitrogen atom. This step may be carried out byusually using trifluoroacetic acid, hydrochloric acid, hydrobromic acidor the like in a halogen-containing solvent such as chloroform ordichloromethane. Alternatively, this step may be carried out by usingtrifluoroacetic acid alone. The reaction temperature is not restricted,and usually a temperature between 0° C. and room temperature isselected. The reaction time may be appropriately selected depending onthe reaction temperature and the like, and usually, the reaction timemay be about 1 to 24 hours.

Formula XX may be produced by the following step using commerciallyavailable asparagine XXV as a starting material:

-   (wherein X₂ and Y₂ represent the same meanings as described above).

Step 1 may be carried out in the same manner as in the reaction betweenFormula XIV and XVII using Formula XXV and XIV. In cases where Z₁ inFormula XIV is chloro or bromo, aqueous sodium hydroxide solution,aqueous potassium hydroxide solution or the like may be used as thebase.

Step 2 may be carried out by reacting Formula XXVI and bromine in abasic solvent such as aqueous sodium hydroxide solution, aqueouspotassium hydroxide solution or the like, for about 1 to 8 hours,although not restricted. The reaction temperature is not restricted, andis usually about room temperature to 100° C. Bromine is usually used inexcess to Formula XXVI. This step may also be carried out by the methoddescribed in J. Org. Chem., 62, 6918(1997) or J. Org. Chem., 49,4272(1984).

Step 3 may be carried out by using thionyl chloride in a solvent such asmethanol at about 0° C. to room temperature. The reaction time is notrestricted, and usually about 1 to 8 hours. The mixing ratio of FormulaXXVII to thionyl chloride is not restricted, and is usually about 1:1 to1:10. The reaction may also be carried out by treating the product withan excess amount of diazomethane or trimethylsilyldiazomethane in asolvent such as methanol at about 0° C. to room temperature.

Step 4 may be carried out by using aqueous sodium hydroxide solution,aqueous potassium hydroxide solution or aqueous potassium carbonatesolution, in an excess amount with respect to Formula XXVIII, in asolvent such as chloroform or dichloromethane at about 0° C. to roomtemperature.

Formula XXIII may be produced by the following steps using commerciallyavailable Formula XXIX.

Step 1 may be carried out in the same manner as in step 2 in the processof producing Formula XX.

Step 2 may be carried out in the same manner as in step 3 in the processof producing Formula XX.

Step 3 may be carried out in the same manner as in step 4 in the processof producing Formula XX.

Among the compounds represented by Formula I, those wherein l=0, m=1,n=1, both R₁ and R₂ are hydrogen atoms, both A and B are nitrogen atoms,C′ and D cooperatively form carbonyl, F is —C(O)—, and R₃, A and R₄cooperatively form Formula II, IV or V, that is, those represented byFormula XXXII, XXXIII and XXXIV:

-   (wherein X₁, Y₁, R₅, R₆, R₇, R₉ and R₁₀ represent the same meanings    as described above) may be produced by hydrolyzing Formula XXXV,    XXXVI and XXXVII:-   (wherein X₁, Y₁, R₅, R₆, R₇, R₉ and R₁₀ represent the same meanings    as described above), respectively, in an alcoholic solvent such as    methanol by a base such as aqueous sodium hydroxide solution or    aqueous barium hydroxide solution. The hydrolysis by the base such    as aqueous sodium hydroxide solution or aqueous barium hydroxide    solution may be carried out, although not restricted, usually at    about 0° C. to room temperature. The amount of the base to be added    is not restricted and usually about 1 to 4 equivalents with respect    to Formula XXXV, XXXVI or XXXVII.

Formula XXXV, XXXVI and XXXII may be produced in the same manner as instep 1 in the process of producing Formula XVII, by using Formula XVIIand Formula XXXVIII, XXXIX and XXXX:

-   (wherein R₆, R₇, R₉ and R₁₀ represent the same meanings as described    above), respectively.

Alternatively, Formula XXXV, XXXI or XXXVII may also be produced byreacting Formula XXXXI, XXXXII or XXXXIII with Formula XVIII or FormulaXIX. This production process may be carried out as in step 1 in theproduction process of Formula XVII.

-   (wherein R₆, R₇, R₉ and R₁₀ represent the same meanings as described    above).

Among the compounds represented by Formula I, those wherein l=0, m=1,n=1, A is a carbon atom, B is a nitrogen atom, C′ and D cooperativelyform carbonyl, F is —CH₂—, R₁ and R₂ are hydrogen atoms, R₃, A and R₄cooperatively form Formula VI, that is, those represented by FormulaXXXXIV:

-   (wherein X₁, X₂, Y₁ and Y₂ represent the same meanings as described    above) may be produced by reacting Formula XVII and Formula XXXXV:-   (wherein X₂ and Y₂ represent the same meanings as described above,    and Z₂ represents a leaving group such as halogen) in a solvent such    as tetrahydrofuran, dichloromethane, dimethylformamide or    acetonitrile, in the presence of a tertiary amine such as    triethylamine or diisopropylamine at, although not restricted,    usually room temperature to refluxing temperature for about 1 to 24    hours, and then hydrolyzing the reaction product under the same    conditions for producing Formula VIII from Formula XI. The mixing    ratio of Formula XVII to Formula XXXXV is about 1:1 to 1:2. The    amount of the tertiary amine to be added is not restricted, and    usually 1 to 4 equivalents with respect to Formula XXXXV.

In cases where the novel spiro derivatives used in the present inventionhave one or more asymmetric carbon atoms, there exist racemic compounds,diasteromers and optical isomers. In the present invention, any of thesemay be used.

The reaction products obtained by the above-described processes may beisolated and purified in the form of a free compound, a salt or asolvate such as hydrate. The salt may be produced by a usualsalt-producing treatment.

Isolation and purification may be carried out by ordinary chemicalprocesses such as extraction, condensation, evaporation,crystallization, filtration, recrystallization and various columnchromatography.

Various isomers may be isolated by conventional methods utilizing thedifferences in the physicochemical properties between the isomers.Optical isomers may be separated by a general optical resolution methodsuch as fractional crystallization or chromatography. Optical isomersmay also be produced by an appropriate optically active compound as thestarting material.

Examples of the pharmaceutically acceptable salts of the compoundsrepresented by Formula I include inorganic salts such as ammonium salt,alkaline metal salts (e.g., sodium salt and potassium salt), alkalineearth metal salts (e.g., calcium salt and magnesium salt); organic saltssuch as dicyclohexylamine salt, N-methyl-D-glucamine salt, ethanolaminesalt, diethanolamine salt, triethanolamine salt, diisopropanolamine saltand tris(hydroxymethyl)aminomethane salt; and lysine- andarginate-addition salts.

Further, various hydrates, solvates and crystalline polymorphs of thecompounds (1) of the present invention and the salts thereof areincluded within the scope of the present invention.

The inhibitory activity of the compound according to the presentinvention against the adhesion of VLA-4 may be determined by using anadhesion-measuring system in which the adhesion between VLA-4-expressingcells such as Ramos cells or Jurkat cells and fibronectin or fibronectinfragment such as a peptide containing CS-1 sequence (Gly Pro Glu Ile LeuAsp Val Pro Ser Thr) (hereinafter referred to as “CS-1 peptide”),immobilized on an immunoplate is measured. Alternatively, abinding-measuring system in which the adhesion between VLA-4 protein andfibronectin or fibronectin fragment such as CS-1 peptide, immobilized onan immunoplate may be used. In the present invention, it is preferred toevaluate the inhibitory activity of a compound using a binding-measuringsystem in which adhesion between a chimera protein ofVLA-4-immunoglobulin (VLA-4-IgG chimera protein) and CS-1 peptide(Japanese Patent Application No. H9-234544), but the method is notrestricted thereto. The “VLA-4-IgG chimera protein” herein means theheterodimer complex of the chimera protein between α4 of VLA-4 andimmunoglobulin (hereinafter referred to as “VLAα4-IgG chimera protein”)and a chimera protein between β1 of VLA-4 and immunoglobulin(hereinafter referred to as “VLAβ1-IgG chimera protein”). As theimmunoglobulin, although heavy chain or light chain of IgG, IgM or thelike may be used, IgG1 heavy chain is used in the present invention.When testing the inhibitory effect of a compound, it is preferred to mixVLA-4-IgG chimera protein and the test compound previously.

Since the compounds according to the present invention have inhibitoryactivities against adhesion of VLA-4, and so inhibit accumulation ofleukocytes at the inflammatory site, they may be used as therapeuticdrugs against chronic inflammatory diseases. Examples of the chronicinflammatory diseases include allergic inflammatory diseases such asbronchial asthma, atopic dermatitis and allergic rhinitis, hepatitis,nephritis, autoimmune diseases such as chronic rheumatoid arthritis andmultiple sclerosis, graft rejections after organ transplantation, type Idiabetes, Crohn's disease and ulcerative colitis. In addition to these,they may be used as therapeutic drugs for the prevention ofpostoperative restenosis, arteriosclerosis and the like.

When using the compound of the present invention as a therapeutic drugagainst the above-mentioned diseases, the compound represented byFormula I or a base addition salt thereof may be administered as it isin the form of powder, or may be administered as a medical compositionin the form of an appropriate formulation, orally or parenterally (e.g.,percutaneous administration, intravenous administration, rectaladministration and inhalation) to mammals.

Examples of the formulation for administration include tablets, powders,balls, capsules, granules, syrups, liquids, injection solutions,emulsions, suspensions and suppositories. These formulations may beprepared by the methods which per se are known, and contain variouscarriers usually used in the field of formulation. Examples thereofinclude vehicles, lubricants, binders and disintegrators for solidformulations; and solvents, solubilizers, suspending agents and soothingagents for liquid formulations. Additives such as antiseptics,antioxidants, coloring agents, sweeteners, absorbents, and wettingagents may be used.

Examples of the vehicles include lactose, D-mannitol, starch, sucrose,corn starch, crystalline cellulose and light anhydrous silicic acid.Examples of the lubricants include magnesium stearate, calcium stearate,talc and colloidal silica. Examples of the binders include crystallinecellulose, saccharose, D-mannitol, dextrin, hydroxypropyl cellulose,hydroxypropylmethyl cellulose, polyvinylpyrrolidone, starch, sucrose,gelatin, methyl cellulose and sodium carboxymethyl cellulose. Examplesof the disintegrators include starch, carboxymethyl cellulose, calciumcarboxymethyl cellulose, cross carmelose sodium, sodium carboxymethylstarch and L-hydroxypropyl cellulose. Examples of the solvents includewater for injection, alcohol, propylene glycol, Macrogol, sesame oil andcorn oil. Examples of the solubilizers include polyethylene glycol,propylene glycol, D-mannitol, benzyl benzoate, ethanol, cholesterol,triethanolamine, sodium carbonate and sodium citrate. Examples of thesuspending agents include surfactants such as stearyl triethanolamine,sodium lauryl sulfate, laurylamino propionate, lecithin, benzalkoniumchloride, benzethonium chloride and glycerin monostearate, andhydrophilic macromolecules such as polyvinyl alcohol, polyvinylpyrrolidone, methyl cellulose, hydroxymethyl cellulose, hydroxyethylcellulose and hydroxypropyl cellulose. Examples of the isotonic agentsinclude glucose, sodium chloride, D-sorbitol and D-mannitol. Examples ofthe buffering agents include buffering solutions containing a phosphoricacid salt, acetic acid salt, carbonic acid salt or citric acid salt. Anexample of the soothing agents is benzylalcohol. Examples of theantiseptics include p-oxybenzoic acid esters, chlorobutanol,benzylalcohol, phenetyl alcohol, dehydroacetic acid and sorbic acid.Examples of the antioxidants include sulfurous acid salts and ascorbicacid.

The effective dose and the number of times of administration of thecompounds represented by Formula I and pharmaceutically acceptable saltsthereof differ depending on the administration form, age and bodyweightof the patient, the type and severity of the disease to be treated, andusually, 1 to 1000 mg, preferably 1 to 300 mg of the compound may beadministered once or in several times per day per adult.

The above-mentioned formulations may contain one or more other effectivecomponents for therapy of other disease(s). Examples thereof includesteroid drugs, nonsteroidal anti-inflammatory drug, lipoxygenaseinhibitors, leucotriene inhibitors, bronchodilators, thromboxanesynthesis inhibitors, thromboxane antagonists, histamine antagonists,histamine release inhibitors, platelet activating factor (PAF)inhibitors, serotonin antagonist, adenosine receptor antagonists,adrenalin β receptor stimulators, immunosuppressors andimmunomodulators.

The effect of the present invention will now be described concretely byway of examples thereof. It should be noted that the present inventionis not restricted to the examples.

EXAMPLE 1 Methyl2-((t-butoxy)carbonylamino)-3-((2,4,8-triaza-1-oxo-4-phenyl-2-benzylspiro[4.5]dec-8-yl)carbonylamino)propionate(1)

Under argon atmosphere, 672 mg of methyl3-amino-2-((t-butoxy)carbonylamino)propionate was dissolved in 15 ml ofacetonitrile and 15 ml of dichloromethane, and then 414 mg of saturatedsodium hydrogen carbonate and 746 mg of chloroformic acid p-nitrophenylester were added thereto while cooling the mixture in ice, followed bystirring the resulting mixture at room temperature for 2.5 hours. To thereaction mixture, 1.385 g of2,4,8-triaza-4-phenyl-2-benzylspiro[4.5]decane-1-one and 2.1 ml oftriethylamine were added, and the resulting mixture was stirred at roomtemperature for 24 hours. After concentrating the reaction mixture,saturated aqueous sodium hydrogen carbonate solution was added, and theresulting mixture was extracted with chloroform. The organic layers werecombined, washed with 0.1N hydrochloric acid and with saturated saline,dried over anhydrous sodium sulfate and concentrated. The residue waspurified by column chromatography (dichloromethane/methanol=80:1) toobtain 1.275 g of methyl2-((t-butoxy)carbonylamino)-3-((2,4,8-triaza-1-oxo-4-phenyl-2-benzylspiro[4.5]dec-8-yl)carbonylamino)propionate (yield: 73%).

LR-MS(m/z): 565(M+) NMR(300 MHz,CDCl3, δ ppm):1.43(9H,s),1.62-1.75(2H,m),2.47-2.62(2H,m),3.53-3.80(3H,m),3.74(3H,s),3.81-3.98(2H,m),4.32-4.42(1H,m),4.55(2H,s),4.60(2H,s), 5.42(1H,brs),5.93(1H,brs),6.63-6.71(2H,m),6.78-6.84(1H,m),7.20-7.40(7H,m)

EXAMPLE 2 Methyl2-amino-3-((2,4,8-triaza-1-oxo-4-phenyl-2-benzylspiro[4.5]dec-8-yl)carbonylamino)propionate(2)

In 2 ml of dichloromethane, 45 mg of methyl2-((t-butoxy)carbonylamino)-3-((2,4,8-triaza-1-oxo-4-phenyl-2-benzylspiro[4.5]dec-8-yl)carbonylamino)propionatewas dissolved, and 1 ml of trifluoroacetic acid was added thereto,followed by stirring the resulting mixture at room temperature for 2hours. After concentrating the reaction mixture, saturated aqueoussodium hydrogen carbonate solution was added to the residue, and theresulting mixture was extracted with chloroform. Organic layers werecombined, washed with saturated saline, dried over anhydrous sodiumsulfate and concentrated to obtain 41 mg of methyl2-amino-3-((2,4,8-triaza-1-oxo-4-phenyl-2-benzylspiro[4.5]dec-8-yl)carbonylamino)propionate.

NMR(300 MHz,CDCl₃, δ ppm):1.72-1.85(2H,m),2.36-2.42(2H,m),3.40-4.20(6H,m),3.80(3H,s),4.15-4.21(1H,m),4.56(2H,s),4.58(2H,s),4.96(1H,brs),6.64-6.70(2H,m),6.83-6.90(1H,m),7.15-7.60(7H,m)

EXAMPLE 33-((2,4,8-triaza-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)-2-((2,6-dichlorophenyl)carbonylamino)propanoicacid (3)

In 3.0 ml of DMF, methyl3-amino-2-((2,6-dichlorophenyl)carbonylamino)propionate (175 mg, 0.601mmol) was dissolved, and2,4,8-triaza-1-oxo-4-phenylspiro[4.5]decane-8-carbonyl chloride (177 mg,0.601 mmol) and triethylamine (250 μl, 1.80 mmol) were added thereto,followed by stirring the resulting mixture at room temperature for 6hours. After adding water to the reaction solution, precipitated solidswere recovered by filtration and washed with water. The solids weredissolved in 3 ml of methanol, and 1 ml of 1N aqueous sodium hydroxidesolution was added thereto, followed by stirring the mixture at roomtemperature for 2 hours. To the reaction solution, 1N hydrochloric acidwas added, and precipitated solids were recovered by filtration. Theobtained solids were purified by column chromatography (ethylacetate/methanol=1/1) to obtain 122 mg of3-((2,4,8-triaza-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)-2-((2,6-dichlorophenyl)carbonylamino)propanoicacid (yield: 38%).

LR-MS(m/z): 533(M⁺) IR(KBr): 3422, 2921, 1708, 1656, 1543, 1432, 1376,1254 cm⁻¹ NMR(300 MHz,CD₃OD, δ ppm): 1.68-1.80 (m, 2H), 2.50-2.64 (m,2H), 3.55-3.80 (m, 4H), 3.90-4.05 (m, 2H), 4.75 (s, 2H), 4.80-4.90 (m,1H), 6.80-6.90 (m, 3H), 7.23-7.35 (m, 2H), 7.38-7.50 (m, 3H). HR-MS:C₂₄H₂₅Cl₂N₅O₅ Calcd.: 532.1154. Found: 532.1140. [α]²⁰ _(D): −10.5°(c=0.30,MeOH)

EXAMPLE 42-((2,6-diemthoxyphenyl)carbonylamino)-3-((2,4,8-triaza-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)propanoicacid (4)

In 10 ml of dichloromethane, 199.2 mg of methyl2-((t-butoxy)carbonylamino)-3-((2,4,8-triaza-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)propionatewas dissolved, and 2 ml of trifluoroacetic acid was added thereto,followed by stirring the resulting mixture at room temperature for 6hours. After concentrating the reaction solution, the residue wasdissolved in 2 ml of dimethylformamide, and then 187 mg of BOP, 70 mg of2,6-dimethoxybenzoic acid and 265 μl of diisopropylamine were addedthereto, followed by stirring the resulting mixture under argonatmosphere at room temperature for 2 hours. To the reaction mixture, 1Nhydrochloric acid was added, and the resulting mixture was extractedwith chloroform. Organic layers were combined, washed with saturatedaqueous sodium hydrogen carbonate solution and with saturated saline,dried over anhydrous sodium sulfate, and concentrated. The residue wasdissolved in 10 ml of methanol, and then 1 ml of 1N aqueous sodiumhydroxide solution was added thereto, followed by stirring the resultingmixture at room temperature for 3 hours. To the reaction mixture, 0.1Nhydrochloric acid was added, and the resulting mixture was extractedwith chloroform. Organic phases were combined, washed with saturatedsaline, dried over anhydrous sodium sulfate, and concentrated. Theresidue was purified by column chromatography (DIOL, ethylacetate/methanol=20:1) to obtain 44.6 mg of2-((2,6-diemthoxyphenyl)carbonylamino)-3-((2,4,8-triaza-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)propanoicacid (yield: 20%).

LR-MS(m/z): 524(M⁺−H) IR(KBr):3408,2927,1656,1602,1542,1468,1432,1381,1262,1196,800,749 cm⁻¹ NMR(300MHz,CDCl₃, δ ppm): 1.68-1.80(2H,m),2.41-2.61(2H,m),3.45-3.57(1H,m),3.62-3.74(2H,m),3.79(6H,s),3.82-3.97(3H,m),4.52-4.57(1H,m),4.70(2H,s),6.18(1H,brs),6.53-6.58(2H,m),6.71-6.78(2H,m),6.83-6.91 (1H,m),7.11 (1H,brs),7.21-7.32(2H,m),7.43-7.48(1H,m) HR-MS:C₂₆H₃₀N₅O₇ Calcd.: 524.2145. Found: 524.2173. [α]²⁰ _(D): −10.0°(c=0.07,MeOH)

EXAMPLE 52-((2,6-dimethoxyphenyl)carbonylamino)-3-((2,4,8-triaza-1-oxo-4-phenyl-2-benzylspiro[4.5]dec-8-yl)carbonylamino)propanoicacid (5)

In 5 ml of acetonitrile, 120 mg of methyl3-amino-2-((2,6-dichlorophenyl)carbonylamino)propionate was dissolved,and 55 mg of saturated sodium hydrogen carbonate and 83 mg ofchloroformic acid p-nitrophenyl ester were added thereto, followed bystirring the resulting mixture at room temperature for 2 hours. To thereaction mixture, 139 mg of2,4,8-triaza-4-phenyl-2-benzylspiro[4.5]decane-1-one and 143 μl oftriethylamine were added, and the resulting mixture was stirred at roomtemperature for 3 hours. After concentrating the reaction mixture,saturated aqueous sodium hydrogen carbonate solution was added, and theresulting mixture was extracted with ethyl acetate. Organic layers werecombined, washed with 1N hydrochloric acid and with saturated saline,dried over anhydrous sodium sulfate, and concentrated. The residue wasdissolved in 2 ml of methanol, and then 0.5 ml of 1N aqueous sodiumhydroxide solution was added thereto, followed by stirring the resultingmixture at room temperature for 4 hours. To the reaction mixture, 0.1Nhydrochloric acid was added, and the resulting mixture was extractedwith chloroform. Organic phases were combined, washed with saturatedsaline, dried over anhydrous sodium sulfate, and concentrated. Theresidue was purified by column chromatography (DIOL, ethylacetate/cyclohexane=2:1) to obtain 18 mg of2-((2,6-dimethoxyphenyl)carbonylamino)-3-((2,4,8-triaza-1-oxo-4-phenyl-2-benzylspiro[4.5]dec-8-yl)carbonylamino)propanoicacid (yield: 7%).

LR-MS(m/z): 622(M⁺−H) IR(KBr):3385,2934,2841,1709,1637,1599,1534,1474,1434,1377,1254,1111,753 cm⁻¹NMR(300 MHz,CD₃OD, δ ppm):1.64-1.73(2H,m),2.43-2.57(2H,m),3.60-3.77(4H,m),3.88-3.98(2H,m),3.82-3.97(3H,m),4.41(2H,s),4.79-4.83(1H,m),4.86(2H,s),6.75-6.82(3H,m),7.18-7.25(2H,m),7.24-7.44(8H,m)HR-MS: C₃₁H₃₀Cl₂N₅O₅ Calcd.: 622.1624. Found: 622.1669. [α]²⁰ _(D):−11.6° (c=0.11,MeOH)

EXAMPLE 6 Methyl2-((2,6-dimethoxyphenyl)carbonylamino)-3-((2,4,8-triaza-1-oxo-4-phenyl-2-benzylspiro[4.5]dec-8-yl)carbonylamino)propionate(6)

In 2 ml of dichloromethane, 49 mg of methyl2-((t-butoxy)carbonylamino)-3-((2,4,8-triaza-1-oxo-4-phenyl-2-benzylspiro[4.5]dec-8-yl)carbonylamino)propionatewas dissolved, and 1 ml of trifluoroacetic acid was added thereto,followed by stirring the resulting mixture at room temperature for 1.5hours. After concentrating the reaction solution, 3 ml ofdichloromethane, 44 mg of BOP, 18 mg of 2,6-dimethoxybenzoic acid and 61μl of diisopropylamine were added to the residue, and the resultingmixture was stirred at room temperature under argon atmosphere for 3hours. To the reaction mixture, 1N hydrochloric acid was added, and theresulting mixture was extracted with chloroform. Organic layers werecombined, washed with saturated sodium hydrogen carbonate solution andwith saturated saline, dried over anhydrous sodium sulfate, andconcentrated. The residue was purified by column chromatography(dichloromethane/methanol=30:1) to obtain 22 mg of methyl2-((2,6-dimethoxyphenyl)carbonylamino)-3-((2,4,8-triaza-1-oxo-4-phenyl-2-benzylspiro[4.5]dec-8-yl)carbonylamino)propionate (yield: 40%).

LR-MS(m/z): 629(M⁺) IR(KBr): 2934, 1743, 1698, 1653, 1598, 1525, 1472,1370, 1254, 1112, 750 cm⁻¹ NMR(300 MHz,CDCl₃, δ ppm):1.63-1.77(2H,m),2.50-2.64(2H,m),3.62-4.06(6H,m),3.80(6H,s),3.81(3H,s),4.56(2H,s),4.60(2H,s),4.85-4.95(1H, m),5.68(1H,brs),6.52-6.58(2H,m),6.64-6.73(2H,m),6.79-6.84(2H,m),7.18-7.40(7H,m)[α]_(D) ²⁰: −24.0° (c=0.05, MeOH)

EXAMPLE 72-((2,6-dimethoxyphenyl)carbonylamino)-3-((2,4,8-triaza-1-oxo-4-phenyl-2-benzylspiro[4.5]dec-8-yl)carbonylamino)propanoicacid (7)

In 1 ml of methanol, 19 mg of methyl2-((2,6-dimethoxyphenyl)carbonylamino)-3-((2,4,8-triaza-1-oxo-4-phenyl-2-benzylspiro[4.5]dec-8-yl)carbonylamino)propionatewas dissolved, and 0.15 ml of 1N aqueous sodium hydroxide solution wasadded thereto, followed by stirring the resulting mixture at roomtemperature for 2 hours. To the reaction solution, 0.1N hydrochloricacid was added, and the resulting mixture was extracted with chloroform.Organic phases were combined, washed with saturated saline, dried overanhydrous sodium sulfate, and concentrated. The residue wasreprecipitated from chloroform/n-hexane to obtain 16.5 mg of2-((2,6-dimethoxyphenyl)carbonylamino)-3-((2,4,8-triaza-1-oxo-4-phenyl-2-benzylspiro[4.5]dec-8-yl)carbonylamino)propanoicacid (yield: 89%).

LR-MS(m/z): 614(M⁺−H) IR(KBr): 3397,2931, 1700, 1653, 1599, 1525, 1510,1473, 1255, 1112, 749 cm⁻¹ NMR(300 MHz,CDCl₃, δ ppm):1.67-1.80(2H,m),2.42-2.65(2H,m),3.40-3.50(1H,m),3.73-4.00(5H,m),3.78(6H,s),4.44-4.51(1H,m),4.57(2H,s),4.62(2H,s),5.99(1H,brs),6.53-6.58(2H,m),6.64-6.77(2H,m),6.82-6.88(1H,m),7.20-7.40(7H,m),7.52-7.59(1H,m)HR-MS: C₃₃H₃₇N₅O₇ Calcd.: 614.2615. Found: 614.2587. [α]_(D) ²⁰: −5.3°(c=0.04, MeOH)

EXAMPLE 8 Methyl2-((2,6-difluorophenyl)carbonylamino)-3-((2,4,8-triaza-1-oxo-4-phenyl-2-benzylspiro[4.5]dec-8-yl)carbonylamino)propionate(8)

In 2 ml of dichloromethane, 78 mg of methyl2-((t-butoxy)carbonylamino)-3-((2,4,8-triaza-1-oxo-4-phenyl-2-benzylspiro[4.5]dec-8-yl)carbonylamino)propionatewas dissolved, and 1 ml of trifluoroacetic acid was added thereto,followed by stirring the resulting mixture at room temperature for 1hour. After concentrating the reaction solution, 2 ml ofdichloromethane, 98 μl of triethylamine, and 26 μl of2,6-difluorobenzoyl chloride were added to the residue, and theresulting mixture was stirred at room temperature for 12.5 hours. To thereaction mixture, saturated aqueous sodium hydrogen carbonate solutionwas added, and the resulting mixture was extracted with chloroform.Organic layers were combined, washed with 0.1N hydrochloric acid andwith saturated saline, dried over anhydrous sodium sulfate, andconcentrated. The residue was purified by column chromatography(dichloromethane/methanol=50:1) to obtain 24 mg of methyl2-((2,6-difluorophenyl)carbonylamino)-3-((2,4,8-triaza-1-oxo-4-phenyl-2-benzylspiro[4.5]dec-8-yl)carbonylamino)propionate (yield: 28%).

LR-MS(m/z): 605(M⁺) IR(KBr): 3064, 3032, 2907, 1748, 1697, 1625, 1601,1540, 1468, 1371, 1260, 1208, 1161, 1122, 1010, 910, 795 cm⁻¹ NMR(300MHz,CDCl₃, δ ppm): 1.64-1.80(2H,m),2.51-2.63(2H,m),3.68-3.98(6H,m),3.79(3H,s),4.58(2H,s),4.61(2H,s),4.79-4.84(1H,m),5.21(1H,brs),6.65-6.72(2H,m),6.78-6.84(1H,m),6.89-6.99(2H,m),7.20-7.40(7H,m),7.78(1H,m)[α]_(D) ²⁰: −18.2° (c=0.11, MeOH)

EXAMPLE 92-((2,6-difluorophenyl)carbonylamino)-3-((2,4,8-triaza-1-oxo-4-phenyl-2-benzylspiro[4.5]dec-8-yl)carbonylamino)propanoicacid (9)

In 1 ml of methanol, 19 mg of methyl2-((2,6-difluorophenyl)carbonylamino)-3-((2,4,8-triaza-1-oxo-4-phenyl-2-benzylspiro[4.5]dec-8-yl)carbonylamino)propionatewas dissolved, and 0.2 ml of 1N aqueous sodium hydroxide solution wasadded thereto, followed by stirring the resulting mixture at roomtemperature for 1 hour. To the reaction solution, 0.1N hydrochloric acidwas added, and the resulting mixture was extracted with chloroform.Organic phases were combined, washed with saturated saline, dried overanhydrous sodium sulfate, and concentrated. The residue wasreprecipitated from chloroform/n-hexane to obtain 15.1 mg of2-((2,6-difluorophenyl)carbonylamino)-3-((2,4,8-triaza-1-oxo-4-phenyl-2-benzylspiro[4.5]dec-8-yl)carbonylamino)propanoicacid (yield: 82%).

LR-MS(m/z): 590(M⁺−H) IR(KBr): 3395, 2927, 1674, 1626, 1601, 1539, 1468,1379, 1262, 1009, 749 cm⁻¹ NMR(300 MHz,CD₃OD, δ ppm):1.71-1.82(2H,m),2.43-2.61(2H,m),3.43-3.54(1H,m),3.73-4.00(5H,m),4.45-4.53(1H,m),4.57(2H,s),4.60(2H,s),5.85(1H,brs),6.66-6.75(2H,m),6.81-6.89(1H,m),6.90-7.00(2H,m),7.20-7.42(7H,m),8.02(1H,m)HR-MS: C₃₁H₃₁F₂N₅O₅ Calcd.: 590.2215. Found: 590.2209. [α]_(D) ²⁰: −73.5(c=0.02, MeOH)

EXAMPLE 10 Methyl2-((2-bromo-6-methylphenyl)carbonylamino)-3-((2,4,8-triaza-1-oxo-4-phenyl-2-benzylspiro[4.5]dec-8-yl)carbonylamino)propionate(10)

In 2 ml of dichloromethane, 158 mg of methyl2-((t-butoxy)carbonylamino)-3-((2,4,8-triaza-1-oxo-4-phenyl-2-benzylspiro[4.5]dec-8-yl)carbonylamino)propionatewas dissolved, and 1 ml of trifluoroacetic acid was added thereto,followed by stirring the resulting mixture at room temperature for 2hours. After concentrating the reaction solution, the residue wasdissolved in 5 ml of dichloromethane, and then 50 mg of BOP, 67 mg of2-bromo-6-methylbenzoic acid and 195 μl of diisopropylamine were addedthereto, followed by stirring the resulting mixture at room temperatureunder argon atmosphere for 20 hours. To the reaction mixture, 1Nhydrochloric acid was added, and the resulting mixture was extractedwith chloroform. Organic layers were combined, washed with saturatedaqueous sodium hydrogen carbonate solution and with saturated saline,dried over anhydrous sodium sulfate, and concentrated. The residue waspurified by column chromatography (dichloromethane/methanol=60:1) toobtain 163 mg of methyl2-((2-bromo-6-methylphenyl)carbonylamino)-3-((2,4,8-triaza-1-oxo-4-phenyl-2-benzylspiro[4.5]dec-8-yl)carbonylamino)propionate(yield: 88%).

LR-MS(m/z): 661 (M⁺) IR(KBr): 2926, 1748, 1699, 1655, 1601, 1526, 1450,1369, 1258, 1206, 1164, 845, 748 cm⁻¹ NMR(300 MHz,CDCl₃, δ ppm):1.64-1.77(2H,m),2.39(3H,s),2.46-2.61(2H,m),3.65-3.94(6H,m),3.80(3H,s),4.57(2H,s),4.59(2H,s),4.80-4.90(1H,m),5.24(1H,brs),6.64-6.73(2H,m),6.80-6.87(1H,m),7.10-7.40(10H,m)[α]_(D) ²⁰: −4.95° (c=0.22, MeOH)

EXAMPLE 112-((2-bromo-6-methylphenyl)carbonylamino)-3-((2,4,8-triaza-1-oxo-4-phenyl-2-benzylspiro[4.5]dec-8-yl)carbonylamino)propanoicacid (11)

In 4 ml of methanol, 150 mg of methyl2-((2-bromo-6-methylphenyl)carbonylamino)-3-((2,4,8-triaza-1-oxo-4-phenyl-2-benzylspiro[4.5]dec-8-yl)carbonylamino)propionatewas dissolved, and 1.2 ml of 1N aqueous sodium hydroxide solution wasadded thereto, followed by stirring the resulting mixture at roomtemperature for 5 hours. To the reaction solution, 1N hydrochloric acidwas added, and the resulting mixture was extracted with chloroform.Organic phases were combined, washed with saturated saline, dried overanhydrous sodium sulfate, and concentrated. The residue wasreprecipitated from chloroform/n-hexane to obtain 121.8 mg of2-((2-bromo-6-methylphenyl)carbonylamino)-3-((2,4,8-triaza-1-oxo-4-phenyl-2-benzylspiro[4.5]dec-8-yl)carbonylamino)propanoicacid (yield: 82%).

LR-MS(m/z): 646(M⁺−H) IR(KBr): 3395, 2926, 1700, 1654, 1601, 1526, 1452,1378, 1261, 749 cm⁻¹ NMR(300 MHz,CD₃OD, δ ppm):1.62-1.80(2H,m),2.36(3H,s),2.43-2.62(2H,m),3.53-3.62(1H,m),3.72-4.00(5H,m),4.57(2H,s),4.60(2H,s),5.99(1H,brs),6.66-6.72(2H,m),6.81-6.90(1H,m),7.12-7.40(9H,m),7.62-7.70(1H,m)HR-MS: C₃₂H₃₄BrN₅O₅ Calcd.: 646.1665. Found: 646.1701. [α]_(D) ²⁰:−30.5° (c=0.04, MeOH)

EXAMPLE 12 Methyl2-((2,6-dimethylphenyl)carbonylamino)-3-((2,4,8-triaza-1-oxo-4-phenyl-2-benzylspiro[4.5]dec-8-yl)carbonylamino)propanoicacid (12)

In 1 ml of dichloromethane, 50 mg of methyl2-((t-butoxy)carbonylamino)-3-((2,4,8-triaza-1-oxo-4-phenyl-2-benzylspiro[4.5]dec-8-yl)carbonylamino)propionatewas dissolved, and 0.5 ml of trifluoroacetic acid was added thereto,followed by stirring the resulting mixture at room temperature for 1hour. After concentrating the reaction solution, the residue wasdissolved in 1 ml of dichloromethane, and then 50 mg of BOP, 15 mg of2,6-dimethylbenzoic acid and 70 μl of diisopropylamine were addedthereto, followed by stirring the resulting mixture overnight underargon atmosphere at room temperature. To the reaction mixture, 1Nhydrochloric acid was added, and the resulting mixture was extractedwith chloroform. Organic layers were combined, washed with saturatedaqueous sodium hydrogen carbonate solution and with saturated saline,dried over anhydrous sodium sulfate, and concentrated. The residue waspurified by column chromatography (ethyl acetate/n-hexane=2:1) to obtain45.8 mg of methyl2-((2,6-dimethylphenyl)carbonylamino)-3-((2,4,8-triaza-1-oxo-4-phenyl-2-benzylspiro[4.5]dec-8-yl)carbonylamino)propanoicacid (yield: 85%).

LR-MS(m/z): 597(M⁺) IR(KBr):3394,2925,1747,1700,1635,1527,1469,1378,1260,1207,1161,750 cm⁻¹ NMR(300MHz,CDCl₃, δ ppm): 1.62-1.71(2H,m),2.32(6H,s),2.40-2.54(2H,m),3.60-3.90(6H,m),3.78(3H,s),4.54(2H,s),4.58(2H,s),4.76-4.84(2H,m),5.63(1H,brs),6.63-6.70(2H,m),6.80-6.84(1H,m),6.93-7.00(2H,m),7.06-7.17(1H,m),7.18-7.43(7H,m),7.40(1H,brs)[α]_(D) ²⁰: −18.0° (c=0.10, CHCl₃)

EXAMPLE 142-((2,6-dimethylphenyl)carbonylamino)-3-((2,4,8-triaza-1-oxo-4-phenyl-2-benzylspiro[4.5]dec-8-yl)carbonylamino)propanoicacid (13)

In 1 ml of methanol, 44 mg of methyl2-((2,6-dimethylphenyl)carbonylamino)-3-((2,4,8-triaza-1-oxo-4-phenyl-2-benzylspiro[4.5]dec-8-yl)carbonylamino)propionatewas dissolved, and 0.4 ml of 1N aqueous sodium hydroxide solution wasadded thereto, followed by stirring the resulting mixture at roomtemperature for 2 hours. To the reaction solution, 1N hydrochloric acidwas added, and the resulting mixture was extracted with chloroform.Organic phases were combined, washed with saturated saline, dried overanhydrous sodium sulfate, and concentrated. The residue wasreprecipitated from chloroform/n-hexane to obtain 23.1 mg of2-((2,6-dimethylphenyl)carbonylamino)-3-((2,4,8-triaza-1-oxo-4-phenyl-2-benzylspiro[4.5]dec-8-yl)carbonylamino)propanoicacid (yield: 54%).

LR-MS(m/z): 682(M⁺−H) IR(KBr):3384,2925,1698,1637,1601,1534,1468,1381,1262,1203,1163,749 cm−1 NMR(300MHz,CD₃OD, δ ppm): 1.72-1.80(2H,m),2.32(6H,s),2.43-2.62(2H,m),3.42-3.51(2H,m),3.75-4.02(4H,m),4.50-4.53(1H,m),4.56(2H,s),4.61 (2H,s),5.83(1H,brs),6.68-6.71(2H,m),6.83-6.90(1H,m),7.01-7.05(2H,m),7.18-7.40(8H,m),7.88(1H,brs)HR-MS: C₃₃H₃₆N₅O₅ Calcd.: 582.2716. Found: 582.2698. [α]²⁰ _(D): −31.5°(c=0.0,CHCl₃)

EXAMPLE 14 Methyl2-(adamantan-2-ylcarbonylamino)-3-((2,4,8-triaza-1-oxo-4-phenyl-2-benzylspiro[4.5]dec-8-yl)carbonylamino)propionate(14)

In 1 ml of dichloromethane, 45 mg of methyl2-((t-butoxy)carbonylamino)-3-((2,4,8-triaza-1-oxo-4-phenyl-2-benzylspiro[4.5]dec-8-yl)carbonylamino)propionatewas dissolved, and 0.5 ml of trifluoroacetic acid was added thereto,followed by stirring the resulting mixture at room temperature for 2hours. After concentrating the reaction solution, the residue wasdissolved in 2 ml of dichloromethane, and then 45 μl of triethylamineand 25 mg of adamantan-2-carbonyl chloride were added thereto, followedby stirring the resulting mixture overnight at room temperature. To thereaction mixture, saturated aqueous sodium hydrogen carbonate solutionwas added, and the resulting mixture was extracted with chloroform.Organic layers were combined, washed with water and with saturatedsaline, dried over anhydrous sodium sulfate, and concentrated. Theresidue was purified by column chromatography (ethylacetate/n-hexane=1:1) to obtain 33 mg of methyl2-(adamantan-2-ylcarbonylamino)-3-((2,4,8-triaza-1-oxo-4-phenyl-2-benzylspiro[4.5]dec-8-yl)carbonylamino)propionate(yield: 65%).

LR-MS(m/z): 627(M⁺) IR(KBr):3423,2908,2851,1739,1702,1638,1526,1453,1370,1260,1203,1151,748 cm⁻¹NMR(300 MHz,CDCl₃, δ ppm):1.61-1.78(8H,m),1.80-1.84(6H,m),1.97-2.05(3H,m),2.46-2.60(2H,m),3.51-3.98(6H,m),3.77(3H,s),4.52-4.60(2H,m),4.55(2H,s),4.61(2H,s),5.33(1H,brs),6.64-6.68(2H,m),6.80-6.84(1H,m),7.20-7.40(7H,m)[α]²⁰ _(D): −13.0° (c=0.10,CHCl₃)

EXAMPLE 152-(adamantan-2-ylcarbonylamino)-3-((2,4,8-triaza-1-oxo-4-phenyl-2-benzylspiro[4.5]dec-8-yl)carbonylamino)propanoicacid (15)

In 2 ml of methanol, 32.1 mg of methyl2-(adamantan-2-ylcarbonylamino)-3-((2,4,8-triaza-1-oxo-4-phenyl-2-benzylspiro[4.5]dec-8-yl)carbonylamino)propionatewas dissolved, and 0.3 ml of 1N aqueous sodium hydroxide solution wasadded thereto, followed by stirring the resulting mixture at roomtemperature for 1.5 hours. To the reaction solution, 1N hydrochloricacid was added, and the resulting mixture was extracted with chloroform.Organic phases were combined, washed with saturated saline, dried overanhydrous sodium sulfate, and concentrated. The residue wasreprecipitated from chloroform/ether to obtain 13.9 mg of2-(adamantan-2-ylcarbonylamino)-3-((2,4,8-triaza-1-oxo-4-phenyl-2-benzylspiro[4.5]dec-8-yl)carbonylamino)propanoicacid (yield: 44%).

LR-MS(m/z): 612(M⁺−H) IR(KBr):3423,2907,2851,1701,1637,1526,1451,1365,1261,746 cm⁻¹ NMR(300 MHz,CDCl₃,δ ppm):1.60-1.81(8H,m),1.82-1.89(6H,m),2.02-2.09(3H,m),2.40-2.62(2H,m),3.40-3.44(2H,m),3.72-4.02(4H,m),4.23-4.28(1H,m),4.56(2H,s),4.61(2H,s),5.64(1H,brs),6.65-6.70(2H,m),6.82-6.87(1H,m),7.19-7.40(7H,m),8.47(1H,brs)HR-MS: C₃₅H₄₂N₅O₅ Calcd.: 612.3186. Found: 612.3190. [α]²⁰ _(D): −15.0°(c=0.1,MeOH)

EXAMPLE 16 Methyl2-((2,6-dichlorophenyl)carbonylamino)-3-((2,4,8-triaza-2-methyl-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)propionate(16)

In 10 ml of dichloromethane, 103 mg of methyl3-amino-2-((2,6-dichlorophenyl)carbonylamino)propionate was dissolved,and 52.9 mg of saturated sodium hydrogen carbonate and 84.7 mg ofchloroformic acid p-nitrophenyl ester were added thereto under argonatmosphere, followed by stirring the resulting mixture at roomtemperature for 1.5 hours. To the reaction mixture, 130 mg of2,4,8-triaza-2-methyl-4-phenylspiro[4.5]decane-1-one and 123 μl oftriethylamine were added, and the resulting mixture was stirredovernight at room temperature. After concentrating the reaction mixture,saturated aqueous sodium hydrogen carbonate solution was added, and theresulting mixture was extracted with chloroform. Organic layers werecombined, washed with 1N hydrochloric acid and with saturated saline,dried over anhydrous sodium sulfate, and concentrated. The residue waspurified by column chromatography (ethyl acetate/methanol=50:1) toobtain 159.8 mg of methyl2-((2,6-dichlorophenyl)carbonylamino)-3-((2,4,8-triaza-2-methyl-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)propionate(yield: 81%).

LR-MS(m/z): 561 (M⁺) IR(KBr): 3395, 3278, 2952, 2932, 1742, 1703, 1655,1616, 1532, 1520, 1432, 1272, 749 cm⁻¹ NMR(300 MHz,CDCl₃, δ ppm):1.63-1.72(2H,m),2.46-2.60(2H,m),3.00(3H,s),3.63-3.92(2H,m),3.82(3H,s),4.68(2H,s),4.82-4.88(1H,m),5.26(1H,brs),6.73-6.78(2H,m),6.84-6.89(1H,m),7.23-7.34(5H,m),7.40(1H,brs)[α]_(D) ²⁰: −15.4° (c=0.10, MeOH)

EXAMPLE 172-((2,6-dichlorophenyl)carbonylamino)-3-((2,4,8-triaza-2-methyl-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)propanoicacid (17)

In 2 ml of methanol, 140 mg of methyl2-((2,6-dichlorophenyl)carbonylamino)-3-((2,4,8-triaza-2-methyl-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)propionatewas dissolved, and 0.75 ml of 1N sodium hydroxide solution was addedthereto, followed by stirring the resulting mixture overnight at roomtemperature. To the reaction solution, 1N hydrochloric acid was added,and the resulting mixture was extracted with chloroform. Organic phaseswere combined, washed with saturated saline, dried over anhydrous sodiumsulfate, and concentrated. The residue was reprecipitated fromchloroform/n-hexane to obtain 104 mg of2-((2,6-dichlorophenyl)carbonylamino)-3-((2,4,8-triaza-2-methyl-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)propanoicacid (yield: 76%).

LR-MS(m/z): 546(M⁺−H) IR(KBr): 3383, 3064, 2930, 1685, 1603, 1533, 1432,1369, 1265, 1197, 1171, 986, 800, 749 cm⁻¹ NMR(300 MHz,CDCl₃, δ ppm):1.68-1.79(2H,m), 2.42-2.63(2H,m), 3.01(3H,s), 3.45-3.59(1H,m),3.68-4.00(5H,m), 4.53-4.58(1H,m), 4.69(2H,s), 5.98(1H,brs),6.74-6.78(2H,m), 6.85-6.92(1H,m), 7.24-7.35(5H,m), 7.64(1H,brs) [α]_(D)²⁰: −18.5° (c=0.10, CHCl₃)

EXAMPLE 18 Methyl2-((2,6-dichlorophenyl)carbonylamino)-3-((2,4,8-triaza-2-(2-methylpropyl)-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)propionate(18)

In 2.7 ml of dichloromethane, 72.2 mg of methyl2-((t-butoxy)carbonylamino)-3-((2,4,8-triaza-1-oxo-4-phenyl-2-benzylspiro[4.5]dec-8-yl)carbonylamino)propionatewas dissolved, and 0.3 ml of trifluoroacetic acid was added thereto,followed by stirring the resulting mixture at room temperature for 2hours. After concentrating the reaction solution, the residue wasdissolved in 2.7 ml of dichloromethane, and then 150 μl of triethylamineand 50 μl of 2,6-dichlorobenzoyl chloride were added thereto, followedby stirring the resulting mixture at room temperature for 5 hours. Tothe reaction mixture, saturated aqueous sodium hydrogen carbonatesolution was added, and the resulting mixture was extracted withchloroform. Organic layers were combined, washed with 0.1N hydrochloricacid and with saturated saline, dried over anhydrous sodium sulfate, andconcentrated. The residue was purified by column chromatography(chloroform/methanol=50:1) to obtain 58.7 mg of methyl2-((2,6-dichlorophenyl)carbonylamino)-3-((2,4,8-triaza-2-(2-methylpropyl)-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)propionate(yield: 71%).

LR-MS(m/z): 603(M⁺) IR(KBr): 3383, 3063, 2958, 1747, 1694, 1528, 1469,1433, 1375, 1259, 1200, 1169, 1128, 965, 800, 782, 749 cm⁻¹ NMR(300MHz,CDCl₃, δ ppm):0.95(6H,d,J=6.5),1.63-1.74(2H,m),1.95-2.08(1H,m),2.46-2.62(2H,m),3.23(2H,d,J=7.4),3.63-3.94(6H,m),3.83(3H,s),4.68(2H,s),4.82-4.88(1H,m),5.24-5.30(1H,m),6.74-6.79(2H,m),6.83-6.88(1H,m),7.23-7.40(6H,m)[α]_(D) ²⁰: −82.0° (c=0.10, MeOH)

EXAMPLE 192-((2,6-dichlorophenyl)carbonylamino)-3-((2,4,8-triaza-2-(2-methylpropyl)-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)propanoicacid (19)

In 2 ml of methanol, 55.6 mg of methyl2-((2,6-dichlorophenyl)carbonylamino)-3-((2,4,8-triaza-2-(2-methylpropyl)-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)propionatewas dissolved, and 1 ml of 1N aqueous sodium hydroxide solution wasadded thereto, followed by stirring the resulting mixture at roomtemperature for 1.5 hours. To the reaction solution, 1N hydrochloricacid was added, and the resulting mixture was extracted with chloroform.Organic phases were combined, washed with saturated saline, dried overanhydrous sodium sulfate, and concentrated. The residue wasreprecipitated from chloroform/ether to obtain 28.4 mg of2-((2,6-dichlorophenyl)carbonylamino)-3-((2,4,8-triaza-2-(2-methylpropyl)-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)propanoicacid (yield: 52%).

LR-MS(m/z): 588(M⁺−H) IR(KBr): 3393, 2960, 2927, 2876, 1740, 1687, 1603,1531, 1470, 1432, 1378, 1264, 1187, 1170, 967, 801, 749 cm⁻¹ NMR(300MHz,CD₃OD, δ ppm): 0.97(6H,d,J=6.6), 1.63-1.72(2H,m),1.96-2.17(1H,m),2.46-2.60(2H,m),3.25(2H,d,J=7.6),3.53-3.77(4H,m),3.86-3.98(2H,m),4.75(2H,s),4.80-4.85(2H,m),6.80-6.88(3H,m),7.22-7.31(2H,m),7.37-7.43(3H,m)[α]²⁰ _(D)=−15.5° (c=0.10,MeOH) HR-MS: C₁₂H₃₂Cl₂N₅O₅ Calcd.: 588.1812.Found: 588.1812.

EXAMPLE 20 Methyl2-((2-methyl-5-nitrophenyl)carbonylamino)-3-((2,4,8-triaza-2-(2-methylpropyl)-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)propionate(20)

In 3 ml of dichloromethane, 54 mg of methyl2-amino-3-((2,4,8-triaza-2-(2-methylpropyl)-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)propionatewas dissolved, and then 71 mg of BOP, 25 mg of 2-methyl-5-nitrobenzoicacid and 91 μl of diisopropylamine were added, followed by stirring theresulting mixture under argon atmosphere at room temperature for 21hours. Water was added to the reaction mixture and the resulting mixturewas extracted with chloroform. Organic layers were combined, washed withsaturated aqueous sodium hydrogen carbonate solution and with saturatedsaline, dried over anhydrous sodium sulfate, and concentrated. Theresidue was purified by column chromatography(dichloromethane/methanol=10:1) to obtain 60 mg of methyl2-((2-methyl-5-nitrophenyl)carbonylamino)-3-((2,4,8-triaza-2-(2-methylpropyl)-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)propionate(yield: 78%).

LR-MS(m/z): 594(M⁺) IR(KBr): 2958, 2871, 1750, 1698, 1657, 1523, 1469,1438, 1348, 1261, 1206, 1164, 1136,912,740 cm⁻¹ NMR(300 MHz,CDCl₃, δppm):0.96(6H,d,J=6.6),1.64-1.78(2H,m),1.92-2.08(1H,m),2.50-2.61(2H,m),2.61(3H,s),3.23(2H,d,J=7.7),3.65-3.97(6H,m),3.80(3H,s),4.69(2H,s),4.66-4.75(1H,m),5.18(1H,brs),6.71-6.77(2H,m),6.79-6.84(1H,m),7.20-7.30(2H,m),7.33-7.40(1H,m),8.12-8.18(1H,m),8.37-8.40(1H,m),8.56(1H,brs)[α]_(D) ²⁰: −18.7° (c=0.05, MeOH)

EXAMPLE 212-((2-methyl-5-nitrophenyl)carbonylamino)-3-((2,4,8-triaza-2-(2-methylpropyl)-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)propanoicacid (21)

In 2 ml of methanol, 57 mg of methyl2-((2-methyl-5-nitrophenyl)carbonylamino)-3-((2,4,8-triaza-2-(2-methylpropyl)-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)propionatewas dissolved, and 0.5 ml of 1N aqueous sodium hydroxide solution wasadded thereto, followed by stirring the resulting mixture at roomtemperature for 1 hour. To the reaction solution, 1N hydrochloric acidwas added, and the resulting mixture was extracted with chloroform.Organic phases were combined, washed with saturated saline, dried overanhydrous sodium sulfate, and concentrated. The residue wasreprecipitated from chloroform/n-hexane to obtain 39.5 mg of2-((2-methyl-5-nitrophenyl)carbonylamino)-3-((2,4,8-triaza-2-(2-methylpropyl)-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)propanoicacid (yield: 68%).

LR-MS(m/z): 579(M⁺−H) IR(KBr): 3383, 2960, 2929, 1689, 1658, 1523, 1471,1349, 1265, 1166, 835, 745 cm⁻¹ NMR(300 MHz,CD₃OD, δ ppm):0.95(6H,d,J=6.6),1.65-1.80(2H,m),1.92-2.07(1H,m),2.43-2.56(2H,m),2.58(3H,s),3.23(2H,d,J=7.7),3.53-3.65(1H,m),3.66-3.99(5H,m),4.53-4.60(1H,m),4.68(2H,s),5.78(1H,brs),6.70-6.79(2H,m),6.80-6.88(1H,m),7.20-7.30(2H,m),7.33-7.40(1H,m),8.13-8.18(1H,m),8.35-8.38(1H,m),8.77(1H,brs)HR-MS: C₂₉H₃₅N₆O₇ Calcd.: 579.2567. Found: 579.2516. [α]_(D) ²⁰: −40.8°(c=0.04, MeOH)

EXAMPLE 22 Methyl2-((2,6-dichlorophenyl)carbonylamino)-3-((2,4,8-triaza-1-oxo-4-phenyl-2-propylspiro[4.5]dec-8-yl)carbonylamino)propionate(22)

In 3 ml of acetonitrile and 8 ml of dichloromethane, 166 mg of methyl3-amino-2-((2,6-dichlorophenyl)carbonylamino) propionate was dissolved,and 69 mg of saturated sodium hydrogen carbonate and 123 mg ofchloroformic acid p-nitrophenyl ester were added thereto under argonatmosphere, followed by stirring the resulting mixture at roomtemperature for 1 hour. To the reaction mixture, 210 mg and 355 μl oftriethylamine were added, and the reaction mixture was stirred at roomtemperature for 13 hours. After concentrating the reaction mixture,saturated aqueous sodium hydrogen carbonate solution was added, and theresulting mixture was extracted with chloroform. Organic layers werecombined, washed with 0.1N hydrochloric acid and with saturated saline,dried over anhydrous sodium sulfate and concentrated. The residue waspurified by column chromatography (dichloromethane/methanol=60:1) toobtain 232 mg of methyl2-((2,6-dichlorophenyl)carbonylamino)-3-((2,4,8-triaza-1-oxo-4-phenyl-2-propylspiro[4.5]dec-8-yl)carbonylamino)propionate(yield: 77%).

LR-MS(m/z): 589(M⁺) IR(KBr): 2962, 2873, 1748, 1688, 1601, 1530, 1471,1432, 1379, 1258, 1197, 1169, 1127, 989, 801, 781, 748 cm⁻¹ NMR(300MHz,CDCl₃, δ ppm):0.96(3H,t,J=7.1),1.61-1.72(2H,m),2.45-2.60(2H,m),3.40(2H,t,J=7.1),3.61-3.95(4H,m),3.83(3H,s),4.68(2H,s),4.82-4.88(1H,m),5.23(1H,brs),6.73-6.79(2H,m),6.82-6.88(1H,m),7.24-7.35(5H,m),7.39(1H,brs)[α]_(D) ²⁰: −11.3° (c=0.27, MeOH)

EXAMPLE 232-((2,6-dichlorophenyl)carbonylamino)-3-((2,4,8-triaza-1-oxo-4-phenyl-2-propylspiro[4.5]dec-8-yl)carbonylamino)propanoicacid (23)

In 5 ml of methanol, 211 mg of methyl2-((2,6-dichlorophenyl)carbonylamino)-3-((2,4,8-triaza-1-oxo-4-phenyl-2-propylspiro[4.5]dec-8-yl)carbonylamino)propionatewas dissolved, and 2 ml of 1N aqueous sodium hydroxide solution wasadded thereto, followed by stirring the resulting mixture at roomtemperature for 1.5 hours. To the reaction solution, 1N hydrochloricacid was added, and the resulting mixture was extracted with chloroform.Organic phases were combined, washed with saturated saline, dried overanhydrous sodium sulfate, and concentrated. The residue wasreprecipitated from chloroform/n-hexane to obtain 166.3 mg of2-((2,6-dichlorophenyl)carbonylamino)-3-((2,4,8-triaza-1-oxo-4-phenyl-2-propylspiro[4.5]dec-8-yl)carbonylamino)propanoicacid (yield: 77%).

LR-MS(m/z): 574(M⁺−H) IR(KBr): 3383, 2965, 2932, 2874, 1684, 1601, 1533,1475, 1432, 1381, 1261, 1197, 1172, 801, 782, 749 cm⁻¹ NMR(300MHz,CDCl₃, δ ppm):0.97(3H,t,J=7.4),1.60-1.76(4H,m),2.43-2.62(2H,m),3.38-3.43(2H,m),3.60-3.77(3H,m),3.81-3.97(3H,m),4.63-4.72(1H,m),4.70(2H,s),6.03(1H,brs),6.72-6.80(2H,m),6.82-6.90(1H,m),7.22-7.38(5H,m),7.77(1H,brs)HR-MS: C₂₇H₃₀Cl₂N₅O₅ Calcd.: 574.1624. Found: 574.1614. [α]_(D) ²⁰:−15.5° (c=0.04, MeOH)

EXAMPLE 24 Methyl3-((2,4,8-triaza-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)-2-((2,4,8-triaza-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)propionate(24)

Under argon atmosphere, 61 mg of methyl2-amino-3-((2,4,8-triaza-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)propionatewas dissolved in 2 ml of acetonitrile and 3 ml of dichloromethane, andthen 22 mg of saturated sodium hydrogen carbonate and 36 mg ofchloroformic acid p-nitrophenyl ester were added thereto while coolingthe mixture in ice, followed by stirring the resulting mixture at roomtemperature for 1.5 hours. To the reaction mixture, 44 mg of2,4,8-triaza-4-phenylspiro[4.5]decane-1-one and 56 μl of triethylaminewere added, and the resulting mixture was stirred overnight at roomtemperature. After concentrating the reaction mixture, saturated aqueoussodium hydrogen carbonate solution was added, and the resulting mixturewas extracted with ethyl acetate. Organic layers were combined, washedwith 1N hydrochloric acid and with saturated saline, dried overanhydrous sodium sulfate and concentrated. The residue was purified bycolumn chromatography (dichloromethane/methanol=50:1) to obtain 54 mg ofmethyl3-((2,4,8-triaza-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)-2-((2,4,8-triaza-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)propionate(yield: 53%).

LR-MS(m/z): 632(M⁺) IR(KBr): 2927, 2366, 1705, 1623, 1601, 1501,1459,1375, 1256, 1187, 965, 910, 750 cm⁻¹ NMR(300 MHz,CDCl₃, δ ppm):1.60-1.80(6H,m),2.43-2.62(4H,m),3.50-4.00(8H,m),3.71(3H,s),4.43-4.51(1H,m),4.71(4H,s),5.70-5.80(1H,brs),6.70-6.80(4H,m),6.72-6.90(2H,m),7.21-7.33(4H,m)[α]_(D) ²⁰: −19.5° (c=0.11, MeOH)

EXAMPLE 253-((2,4,8-triaza-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)-2-((2,4,8-triaza-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)propanoicacid (25)

In 2 ml of methanol, 36 mg of methyl3-((2,4,8-triaza-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)-2-((2,4,8-triaza-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)propionatewas dissolved, and 0.3 ml of 1N sodium hydroxide solution was addedthereto, followed by stirring the resulting mixture at room temperaturefor 2.5 hours. To the reaction solution, 1N hydrochloric acid was added,and the resulting mixture was extracted with chloroform. Organic phaseswere combined, washed with saturated saline, dried over anhydrous sodiumsulfate, and concentrated. The residue was reprecipitated fromchloroform/ether to obtain 38.7 mg of3-((2,4,8-triaza-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)-2-((2,4,8-triaza-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)propanoicacid (yield: 81%).

LR-MS(m/z): 617(M⁺−H) IR(KBr): 3347, 2929, 1708, 1601, 1533, 1501, 1397,1255, 1189, 966, 751 cm⁻¹ NMR(300 MHz,CD₃OD, δ ppm):1.63-1.80(3H,m),2.10-2.40(4H,m),2.41-2.60(3H,m),3.54-3.75(4H,m),3.76-3.87(2H,m),3.88-4.00(2H,m),4.24-4.35(1H,m),4.73(4H,s),6.65-6.79(4H,m),6.82-6.92(2H,m),7.20-7.30(4H,m)[α]_(D) ²⁰: 26.3° (c=0.04, MeOH) HR-MS: C₃₁H₃₈N₈O₆ Calcd.: 617.2836.Found: 617.2834.

EXAMPLE 26 Methyl2-((4-(2-methylpropanoylamino)-4-phenylpiperidyl)carbonylamino)-3-((2,4,8-triaza-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)propionate(26)

Under argon atmosphere, 41 mg of methyl2-amino-3-((2,4,8-triaza-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)propionatewas dissolved in 2 ml of acetonitrile and 3 ml of dichloromethane, andthen 11 mg of saturated sodium hydrogen carbonate and 18 mg ofchloroformic acid p-nitrophenyl ester were added thereto while coolingthe mixture in ice, followed by stirring the resulting mixture at roomtemperature for 2.5 hours. To the reaction mixture, 24 mg and 28 μl oftriethylamine were added, and the resulting mixture was stirred at roomtemperature for 16 hours. After concentrating the reaction mixture,saturated aqueous sodium hydrogen carbonate solution was added, and theresulting mixture was extracted with ethyl acetate. Organic layers werecombined, washed with 1N hydrochloric acid and with saturated saline,dried over anhydrous sodium sulfate and concentrated. The residue waspurified by column chromatography (dichloromethane/methanol=40:1) toobtain 11 mg of methyl2-((4-(2-methylpropanoylamino)-4-phenylpiperidyl)carbonylamino)-3-((2,4,8-triaza-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)propionate(yield: 19%).

LR-MS(m/z): 647(M⁺) NMR(300 MHz,CDCl₃, δ ppm):1.04-1.18(6H,m),1.63-1.76(2H,m),1.90-2.04(2H,m),2.28-2.41(2H,m),2.51-2.64(2H,m),3.02-3.18(2H,m),3.53-4.00(8H,m),3.78(3H,s),4.43-4.49(1H,m),4.58(2H,s),4.61(2H,s),5.35(1H,brs),5.58(1H,brs)6.64-6.71(2H,m),6.78-6.85(1H,m),6.97-7.02(1H,m),7.16-7.40(11H,m)

EXAMPLE 272-((4-(2-methylpropanoylamino)-4-phenylpiperidyl)carbonylamino)-3-((2,4,8-triaza-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)propanoicacid (27)

In 1 ml of methanol, 11 mg of methyl2-((4-(2-methylpropanoylamino)-4-phenylpiperidyl)carbonylamino)-3-((2,4,8-triaza-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)propionatewas dissolved, and 0.1 ml of 1N aqueous sodium hydroxide solution wasadded thereto, followed by stirring the resulting mixture at roomtemperature for 2.5 hours. To the reaction solution, 1N hydrochloricacid was added, and the resulting mixture was extracted with chloroform.Organic phases were combined, washed with saturated saline, dried overanhydrous sodium sulfate, and concentrated. The residue wasreprecipitated from chloroform/ether to obtain 6.4 mg of2-((4-(2-methylpropanoylamino)-4-phenylpiperidyl)carbonylamino)-3-((2,4,8-triaza-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)propanoicacid (yield: 59%).

LR-MS(m/z): 722(M⁺−H) IR(KBr): 3369, 2966, 2929, 1688, 1628, 1603, 1530,1469, 1381, 1262, 1207, 751 cm⁻¹ NMR(300 MHz,CDCl₃, δ ppm):1.10-1.20(6H,m),1.62-1.80(2H,m),1.93-2.10(2H,m),2.32-2.63(5H,m),3.10-3.30(2H,m),3.55-4.00(8H,m),4.21-4.24(1H,m),4.58(2H,s),4.60(2H,s),5.52(1H,brs),5.61(1H,brs),6.61-6.70(2H,m),6.81-6.88(1H,m),7.20-740(12H,m),8.09(1H,brs)[α]_(D) ²⁰: −5.00° (c=0.02, MeOH) HR-MS: C₄₀H₄₉N₇O₆ Calcd.: 722.3666.Found: 722.3726.

EXAMPLE 28 Methyl2-((4-(2-oxo(3-hydrobenzimidazolyl))piperidyl)carbonylamino)-3-(2,4,8-triaza-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)propionate(28)

Under argon atmosphere, 100.3 mg of methyl3-amino-2-((4-(2-oxo(3-hydrobenzimidazolyl))piperidyl)carbonylamino)propionatewas dissolved in 15 ml of dichloromethane, and 42 mg of saturated sodiumhydrogen carbonate and 67 mg of chloroformic acid p-nitrophenyl esterwere added thereto, followed by stirring the resulting mixture at roomtemperature for 2 hours. To the reaction mixture, 97 mg and 100 μl oftriethylamine were added, and the resulting mixture was stirredovernight at room temperature. After concentrating the reaction mixture,saturated aqueous sodium hydrogen carbonate solution was added, and theresulting mixture was extracted with ethyl acetate. Organic layers werecombined, washed with 1N hydrochloric acid and with saturated saline,dried over anhydrous sodium sulfate and concentrated. The residue waspurified by column chromatography (dichloromethane/methanol=10:1) toobtain 54.7 mg of methyl2-((4-(2-oxo(3-hydrobenzimidazolyl))piperidyl)carbonylamino)-3-(2,4,8-triaza-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)propionate(yield: 32%).

LR-MS(m/z): 618(M⁺) IR(KBr): 3427, 2927, 1704, 1624, 1540, 1381, 1254,1158, 1087, 753 cm⁻¹ NMR(300 MHz,CDCl₃, δ ppm):1.65-1.85(4H,m),2.23-2.40(2H,m),2.49-2.63(2H,m),2.83-2.98(2H,m),3.47-3.51(1H,m),3.60-3.98(5H,m),3.78(3H,s),4.17-4.28(2H,m),4.38-4.52(2H,m),4.75(2H,s),5.40(1H,brs),6.70-6.78(2H,m),6.84-6.92(1H,m),6.97-7.09(4H,m),7.24-7.35(2H,m),8.75(1H,brs)[α]_(D) ²⁰: −6.85° (c=0.10, MeOH)

EXAMPLE 292-((4-(2-oxo(3-hydrobenzimidazolyl))piperidyl)carbonylamino)-3-(2,4,8-triaza-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)propanoicacid (29)

In 2 ml of methanol, 48 mg of methyl2-((4-(2-oxo(3-hydrobenzimidazolyl))piperidyl)carbonylamino)-3-(2,4,8-triaza-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)propionatewas dissolved, and 0.3 ml of 1N sodium hydroxide solution was addedthereto, followed by stirring the resulting mixture overnight at roomtemperature. To the reaction solution, 1N hydrochloric acid was added,and the resulting mixture was extracted with chloroform. Organic phaseswere combined, washed with saturated saline, dried over anhydrous sodiumsulfate, and concentrated. The residue was reprecipitated fromchloroform/ether to obtain 24.8 mg of2-((4-(2-oxo(3-hydrobenzimidazolyl))piperidyl)carbonylamino)-3-(2,4,8-triaza-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)propanoicacid (yield: 53%).

LR-MS(m/z): 603(M⁺−H) IR(KBr): 3383, 2932, 1702, 1622, 1604, 1541, 1488,1379, 1254, 1192, 1162, 1091, 966, 754 cm⁻¹ NMR(300 MHz,CD₃OD, δ ppm):1.60-1.78(4H,m),2.20-2.40(2H,m),2.50-2.62(2H,m),2.86-3.00(2H,m),3.50-3.57(4H,m),3.86-4.00(2H,m),4.17-4.30(2H,m),4.31-4.37(1H,m),4.40-4.52(1H,m),4.73(2H,s),6.75-6.83(3H,m),6.92-7.05(4H,m),7.21-7.30(2H,m) [α]_(D) ²⁰: −14.70 (c=0.01, MeOH)

EXAMPLE 30 Methyl2-((4-(2-oxo(3-hydrobenzimidazolyl))piperidyl)carbonylamino)-3-((2,4,8-triaza-1-oxo-4-phenyl-2-benzylspiro[4.5]dec-8-yl)carbonylamino)propionate(30)

Under argon atmosphere, 45.1 mg of methyl2-amino-3-((2,4,8-triaza-1-oxo-4-phenyl-2-benzylspiro[4.5]dec-8-yl)carbonylamino)propionatewas dissolved in 2 ml of dichloromethane, and then 11.1 mg of saturatedsodium hydrogen carbonate and 21.3 mg of chloroformic acid p-nitrophenylester were added thereto while cooling the mixture in ice, followed bystirring the resulting mixture at room temperature for 2.5 hours. To thereaction mixture, 28.7 mg of 1-(4-piperidyl)-3-hydrobenzimidazol-2-oneand 65 μl of triethylamine were added, and the resulting mixture wasstirred at room temperature for 8 hours. After concentrating thereaction mixture, saturated aqueous sodium hydrogen carbonate solutionwas added, and the resulting mixture was extracted with ethyl acetate.Organic layers were combined, washed with 1N hydrochloric acid and withsaturated saline, dried over anhydrous sodium sulfate and concentrated.The residue was purified by column chromatography(dichloromethane/methanol=20:1) to obtain 38.7 mg of methyl2-((4-(2-oxo(3-hydrobenzimidazolyl))piperidyl)carbonylamino)-3-((2,4,8-triaza-1-oxo-4-phenyl-2-benzylspiro[4.5]dec-8-yl)carbonylamino)propionate(yield: 60%).

LR-MS(m/z): 708(M⁺) IR(KBr): 3387, 2931, 1689, 1623, 1543, 1484, 1379,1267, 1204, 1137, 754 cm⁻¹ NMR(300 MHz,CDCl₃, δ ppm):1.63-1.85(4H,m),2.22-2.38(2H,m),2.52-2.66(2H,m),2.83-2.99(2H,m),3.60-3.88(4H,m),3.77(3H,s),3.91-4.00(1H,m),4.15-4.30(2H,m),4.40-4.53(2H,m),4.57(2H,s),4.61(2H,s),5.30-5.38(1H,m),6.63-6.72(2H,m),6.80-6.86(1H,m),6.90-7.08(6H,m),7.21-7.40(5H,m),8.57(1H,brs)[α]_(D) ²⁰: −124.0° (c=0.10, MeOH)

EXAMPLE 312-((4-(2-oxo(3-hydrobenzimidazolyl))piperidyl)carbonylamino)-3-((2,4,8-triaza-1-oxo-4-phenyl-2-benzylspiro[4.5]dec-8-yl)carbonylamino)propanoicacid (31)

In 1.5 ml of methanol, 36.6 mg of methyl2-((4-(2-oxo(3-hydrobenzimidazolyl))piperidyl)carbonylamino)-3-((2,4,8-triaza-1-oxo-4-phenyl-2-benzylspiro[4.5]dec-8-yl)carbonylamino)propionatewas dissolved, and 1 ml of 1N aqueous sodium hydroxide solution wasadded thereto, followed by stirring the resulting mixture at roomtemperature for 2.5 hours. To the reaction solution, 1N hydrochloricacid was added, and the resulting mixture was extracted with ethylacetate. Organic phases were combined, washed with saturated saline,dried over anhydrous sodium sulfate, and concentrated. The residue wasreprecipitated from chloroform/n-hexane to obtain 33.2 mg of2-((4-(2-oxo(3-hydrobenzimidazolyl))piperidyl)carbonylamino)-3-((2,4,8-triaza-1-oxo-4-phenyl-2-benzylspiro[4.5]dec-8-yl)carbonylamino)propanoicacid (yield: 92%).

LR-MS(m/z): 695(M⁺+H) IR(KBr): 3368, 2927, 2862, 1697, 1631, 1535, 1485,1370, 1263, 1196, 1157, 1086, 1011, 970, 751 cm⁻¹ NMR(300 MHz,CD₃OD, δppm): 1.60-1.80(4H,m),2.20-2.40(2H,m),2.50-2.61(2H,m),2.89-3.01(2H,m),3.60-3.75(4H,m),3.91-4.02(2H,m),4.16-4.30(2H,m),4.30-4.55(2H,m),4.60(2H,s),4.62(2H,s),6.62-6.82(3H,m),6.87-7.07(4H,m),7.20-7.40(7H,m)HR-MS: C₃₇H₄₃N₈O₆ Calcd.: 695.3306. Found: 695.3339. [α]²⁰ _(D): −12.0°(c=0.10,MeOH)

EXAMPLE 32 Methyl2-((4-(2-oxo(3-hydrobenzimidazolyl))piperidyl)carbonylamino)-3-((2,4,8-triaza-2-(2-methylpropyl)-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)propionate(32)

In 2.7 ml of dichloromethane, 173.1 mg of methyl2-((t-butoxy)carbonylamino)-3-((2,4,8-triaza-2-(2-methylpropyl)-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)propionatewas dissolved, and 0.3 ml of trifluoroacetic acid was added thereto. Theresulting mixture was stirred at room temperature for 2 hours, and thenconcentrated. Under argon atmosphere, the residue was dissolved in 2.4ml of acetonitrile, and then 15.4 mg of saturated sodium hydrogencarbonate and 29.5 mg of chloroformic acid p-nitrophenyl ester wereadded thereto, followed by stirring the resulting mixture at roomtemperature for 4 hours. To the reaction mixture, 39.8 mg of1-(4-piperidyl)-3-hydrobenzimidazol-2-one and 85 μl of triethylaminewere added, and the resulting mixture was stirred overnight at roomtemperature. After concentrating the reaction mixture, saturated aqueoussodium hydrogen carbonate solution was added, and the resulting mixturewas extracted with chloroform. Organic layers were combined, washed with0.1N hydrochloric acid and with saturated saline, dried over anhydroussodium sulfate and concentrated. The residue was purified by columnchromatography (chloroform/methanol=30:1) to obtain 55.6 mg of methyl2-((4-(2-oxo(3-hydrobenzimidazolyl))piperidyl)carbonylamino)-3-((2,4,8-triaza-2-(2-methylpropyl)-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)propionate(yield: 59%)

LR-MS(m/z): 674(M⁺) IR(KBr): 3363, 2957, 2869, 1697, 1629, 1536, 1483,1371, 1265, 1200, 1153, 1087, 1011, 966, 752 cm⁻¹ NMR(300 MHz,CDCl₃, δppm):0.95(6H,d,J=6.6),1.60-1.75(2H,m),1.78-1.85(2H,m),1.96-2.08(1H,m),2.22-2.38(2H,m),2.53-2.64(2H,m),2.84-3.00(2H,m),3.23(2H,d,J=7.4),3.60-4.00(6H,m),3.77(3H,s),4.17-4.30(2H,m),4.40-4.55(2H,m),4.70(2H,s),5.26-5.34(1H,m),6.72-6.77(2H,m),6.83-6.90(1H,m),6.98-7.10(5H,m),7.24-7.33(2H,m),8.39(1H,brs)[α]²⁰ _(D): −6.5° (c=0.10,MeOH)

EXAMPLE 332-((4-(2-oxo(3-hydrobenzimidazolyl))piperidyl)carbonylamino)-3-((2,4,8-triaza-2-(2-methylpropyl)-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)propanoicacid (33)

In 1 ml of methanol, 53.2 mg of methyl2-((4-(2-oxo(3-hydrobenzimidazolyl))piperidyl)carbonylamino)-3-((2,4,8-triaza-2-(2-methylpropyl)-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)propionatewas dissolved, and 1 ml of 1N aqueous sodium hydroxide solution wasadded thereto, followed by stirring the resulting mixture at roomtemperature for 2 hours. To the reaction solution, 1N hydrochloric acidwas added, and the resulting mixture was extracted with ethyl acetate.Organic phases were combined, washed with saturated saline, dried overanhydrous sodium sulfate, and concentrated. The residue wasreprecipitated from chloroform/n-hexane to obtain 36.6 mg of2-((4-(2-oxo(3-hydrobenzimidazolyl))piperidyl)carbonylamino)-3-((2,4,8-triaza-2-(2-methylpropyl)-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)propanoicacid (yield: 70%).

LR-MS(m/z): 659(M⁺−H) IR(KBr): 3372, 2959, 2871, 1696, 1634, 1536, 1483,1376, 1266, 1197, 1163, 1090, 1011, 968, 752 cm⁻¹ NMR(300 MHz,CD₃OD, δppm):0.95(6H,d,J=6.6),1.59-1.69(2H,m),1.70-1.80(2H,m),1.99-2.17(1H,m),2.22-2.40(2H,m),2.50-2.63(2H,m),2.90-3.03(2H,m),3.24(2H,d,J=7.7),3.58-3.70(4H,m),3.90-4.02(2H,m),4.18-4.31(2H,m),4.32-4.39(1H,m),4.40-4.53(1H,m),4.76(2H,s),6.80-6.87(3H,m),6.92-7.10(4H,m),7.24-7.33(2H,m)[α]²⁰ _(D): −10.0° (c=0.10,MeOH) HR-MS: C₃₄H₄₅N₈O₆ Calcd.: 661.3462.Found: 661.3450.

EXAMPLE 34 Methyl2-((2,6-dichlorophenyl)amino)carbonylamino)-3-((2,4,8-triaza-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)propionate(34)

In 2 ml of dichloromethane, 100 mg of methyl2-((t-butoxy)carbonylamino)-3-((2,4,8-triaza-1-oxo-4-phenyl-2-benzylspiro[4.5]dec-8-yl)carbonylamino)propionatewas dissolved, and 1 ml of trifluoroacetic acid was added thereto,followed by stirring the resulting mixture at room temperature for 2hours. After concentrating the reaction solution, the residue wasdissolved in 4 ml of dichloromethane, and then 150 μl of triethylamineand 43 mg of 2,6-dichlorobenzene isocyanate were added, followed bystirring the resulting mixture at room temperature for 3 hours.Saturated aqueous sodium hydrogen carbonate solution was added to thereaction mixture, and the resulting mixture was extracted withchloroform. Organic layers were combined, washed with water and withsaturated saline, dried over anhydrous sodium sulfate, and concentratedto obtain 90.7 mg of methyl2-((2,6-dichlorophenyl)amino)carbonylamino)-3-((2,4,8-triaza-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)propionate(yield: 77%).

LR-MS(m/z): 562(M⁺) IR(KBr):3371,2925,2852,1708,1624,1539,1457,1438,1372,1254,1078,782,750 cm⁻¹NMR(300 MHz,CDCl₃, δ ppm):1.68-1.81(2H,m),2.43-2.57(2H,m),3.58-3.87(6H,m),3.75(3H,s),4.52-4.60(1H,m),4.75(2H,s),5.26(1H,brs),6.13(1H,brs),6.25(1H,brs),6.33(1H,brs),6.72-6.78(2H,m),6.85-6.90(1H.m),7.10-7.20(2H,m),7.21-7.39(3H,m)[α]²⁰ _(D): −5.5° (c=0.10,CHCl₃)

EXAMPLE 352-((2,6-dichlorophenyl)amino)carbonylamino)-3-((2,4,8-triaza-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)propanoicacid (35)

In 4 ml of methanol, 90 mg of methyl2-((2,6-dichlorophenyl)amino)carbonylamino)-3-((2,4,8-triaza-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)propionatewas dissolved, and 1.6 ml of 1N aqueous sodium hydroxide solution wasadded thereto, followed by stirring the resulting mixture at roomtemperature for 4 hours. To the reaction solution, 1N hydrochloric acidwas added, and the resulting mixture was extracted with chloroform.Organic phases were combined, washed with saturated saline, dried overanhydrous sodium sulfate, and concentrated. The residue wasreprecipitated from chloroform/ether to obtain 21.0 mg of2-((2,6-dichlorophenyl)amino)carbonylamino)-3-((2,4,8-triaza-1-oxo-4-phenylspiro[4.5]dec-8-yl)carbonylamino)propanoicacid (yield: 52%).

LR-MS(m/z): 547(M⁺−H) IR(KBr):3355,2931,1707,1604,1539,1457,1437,1373,1255,1203,1164,1094,965,781 cm⁻¹NMR(300 MHz,CD₃OD, δ ppm):1.62-1.74(2H.m),2.45-2.60(2H,m),3.55-3.70(4H,m),3.83-3.99(2H,m),4.42-4.50(1H,m),4.71(2H,s), 6.78-6.83(3H,m),7.20-7.30(3H.m),7.38-7.41(2H,m) HR-MS:C₂₄H₂₅Cl₂N₆O₅ Calcd.: 547.1263. Found: 547.1254. [α]²⁰ _(D):−9.5(c=0.10,MeOH)

EXAMPLE 36 Methyl2-((2,6-dichlorophenyl)amino)carbonylamino)-3-((2,4,8-triaza-1-oxo-4-phenyl-2-benzylspiro[4.5]dec-8-yl)carbonylamino)propionate(36)

In 1 ml of dichloromethane, 50 mg of methyl2-((t-butoxy)carbonylamino)-3-((2,4,8-triaza-1-oxo-4-phenyl-2-benzylspiro[4.5]dec-8-yl)carbonylamino)propionate was dissolved, and0.5 ml of trifluoroacetic acid was added thereto, followed by stirringthe resulting mixture at room temperature for 2 hours. Afterconcentrating the reaction solution, the residue was dissolved in 2 mlof dichloromethane, and then 60 μl of triethylamine and 19 mg of2,6-dichlorobenzene isocyanate were added, followed by stirring theresulting mixture overnight at room temperature. Saturated aqueoussodium hydrogen carbonate solution was added to the reaction mixture,and the resulting mixture was extracted with chloroform. Organic layerswere combined, washed with water and with saturated saline, dried overanhydrous sodium sulfate, and concentrated. The residue was purified bycolumn chromatography (ethyl acetate/n-hexane=1:1) to obtain 46.3 mg ofmethyl2-((2,6-dichlorophenyl)amino)carbonylamino)-3-((2,4,8-triaza-1-oxo-4-phenyl-2-benzylspiro[4.5]dec-8-yl)carbonylamino)propionate(yield: 80%).

LR-MS(m/z): 652(M⁺) IR(KBr):3392,2925,1740,1701,1651,1539,1455,1436,1375,1261,1201,1162,1063,75 2cm−1 NMR(300 MHz,CDCl₃, δ ppm):1.65-1.78(2H,m),2.45-2.60(2H,m),3.60-3.90(6H,m),3.78(3H,s),4.52-4.61(1H,m),4.54(2H,s),4.60(2H,s),5.40(1H,brs),6.44(1H,brs),6.63-6.78(3H,m),6.79-6.84(1H,m),7.07-7.12(1H,m),7.20-7.40(9H,m)[α]²⁰ _(D): −7.0° (c=0.10,CHCl₃)

EXAMPLE 372-((2,6-dichlorophenyl)amino)carbonylamino)-3-((2,4,8-triaza-1-oxo-4-phenyl-2-benzylspiro[4.5]dec-8-yl)carbonylamino)propanoicacid (37)

In 2 ml of methanol, 41.2 mg of methyl2-((2,6-dichlorophenyl)amino)carbonylamino)-3-((2,4,8-triaza-1-oxo-4-phenyl-2-benzylspiro[4.5]dec-8-yl)carbonylamino)propionatewas dissolved, and 0.6 ml of 1N aqueous sodium hydroxide solution wasadded thereto, followed by stirring the resulting mixture at roomtemperature for 2 hours. To the reaction solution, 1N hydrochloric acidwas added, and the resulting mixture was extracted with chloroform.Organic phases were combined, washed with saturated saline, dried overanhydrous sodium sulfate, and concentrated. The residue wasreprecipitated from chloroform/ether to obtain 21.0 mg of2-((2,6-dichlorophenyl)amino)carbonylamino)-3-((2,4,8-triaza-1-oxo-4-phenyl-2-benzylspiro[4.5]dec-8-yl)carbonylamino)propanoicacid (yield: 52%).

LR-MS(m/z): 637(M+−H) IR(KBr):3418,2926,1724,1700,1624,1539,1472,1377,1265,1194,1075,1025,786,750 cm⁻¹NMR(300 MHz,CDCl₃, δ ppm):1.64-1.78(2H,m),2.40-2.60(2H,m),3.44-3.55(2H,m),3.63-3.92(4H,m),4.30-4.40(1H,m),4.54(2H,s),4.60(2H,s),5.19(1H,brs),5.73(1H,brs),6.63-6.70(2H,m),6.79-6.89(2H,m),7.10-7.45(9H,m)[α]²⁰ _(D): −6.0° (c=0.10,MeOH)

EXAMPLE 382-((2,6-dichlorophenyl)carbonylamino)-4-oxo-4-(2,4,8-triaza-1-oxo-4-phenylspiro[4.5]dec-8-yl)butanoicacid (38)

In 2 ml of DMF, 150 mg of methyl2-amino-4-oxo-4-(2,4,8-triaza-1-oxo-4-phenylspiro[4.5]dec-8-yl)butanoatewas dissolved, and 97 μl of 2,6-dichlorobenzoyl chloride and 192 μl oftriethylamine were added thereto, followed by stirring the resultingmixture at room temperature for 14 hours. Saturated aqueous sodiumhydrogen carbonate solution was added to the reaction mixture, and theresulting mixture was extracted with ethyl acetate. Organic phases werecombined, washed with 0.1N hydrochloric acid and with saturated saline,dried over anhydrous sodium sulfate, and concentrated. The residue wasdissolved in 6 ml of methanol, and 2 ml of 1N aqueous sodium hydroxidesolution was added thereto, followed by stirring the resulting mixturefor 1.5 hours. To the reaction solution, 1N hydrochloric acid was addedand the resulting mixture was extracted with ethyl acetate. Organicphases were combined, washed with saturated saline, dried over anhydroussodium sulfate, and concentrated. The residue was purified by columnchromatography (DIOL, ethyl acetate/methanol=10/1) to obtain 270 mg of2-((2,6-dichlorophenyl)carbonylamino)-4-oxo-4-(2,4,8-triaza-1-oxo-4-phenylspiro[4.5]dec-8-yl)butanoicacid (yield: 80%).

LR-MS(m/z): 517(M⁺−H) IR(KBr):3408,2926,1709,1638,1601,1500,1459,1432,1382,1235,1195 cm⁻¹ NMR(300MHz,CDCl₃, δ ppm):2.35-2.65(2H,m),3.00-3.25(2H,m),3.40-3.50(1H,m),3.80-4.00(2H,m),4.30-4.45(1H,m),4.71(2H,s),5.00-5.07(1H,m),6.80-6.90(3H,m),7.20-7.30(2H,m),7.50-7.65(3H,m)HR-MS: C₂₄H₂₄Cl₂N₄O₅ Calcd.: 517.1046. Found: 517.1092. [α]²⁰ _(D):+3.37° (c=0.31,MeOH)

EXAMPLE 39t-butyl-2-((2,6-dichlorophenyl)carbonylamino)-5-oxo-5-(2,4,8-triaza-1-oxo-4-phenylspiro[4.5]dec-8-yl)pentanoate(39)

In 2 ml of DMF, 339 mg of t-butyl2-amino-5-oxo-5-(2,4,8-triaza-1-oxo-4-phenylspiro[4.5]dec-8-yl)pentanoatewas dissolved, and 197 μl of 2,6-dichlorobenzoyl chloride and 385 μl oftriethylamine were added thereto, followed by stirring the resultingmixture at room temperature for 3 hours. Saturated aqueous sodiumhydrogen carbonate solution was added to the reaction mixture, and theresulting mixture was extracted with ethyl acetate. Organic phases werecombined, washed with 0.1N hydrochloric acid and with saturated saline,dried over anhydrous sodium sulfate, and concentrated. The residue waspurified by column chromatography (ethyl acetate/n-hexane=6/1) to obtain468 mg of t-butyl2-((2,6-dichlorophenyl)carbonylamino)-5-oxo-5-(2,4,8-triaza-1-oxo-4-phenylspiro[4.5]dec-8-yl)pentanoate(yield: 86%).

LR-MS(m/z): 588(M⁺) IR(KBr): 3259, 3066, 2979, 2932, 1717, 1627, 1502,1432, 1367, 1299, 1249, 1155, 1088, 1046, 964, 846, 801, 749 cm⁻¹NMR(300 MHz,CDCl₃, δ ppm): 1.50(9H,s),1.70-1.87(2H,m),2.00-2.20(1H,m),2.30-2.76(5H,m),3.35-3.45(1H,m),3.73-3.98(2H,m),4.46-4.61(1H,m),4.68-4.82(1H,m),4.76(2H,s),6.60(1H,brs),6.74-6.97(4H,m),7.21-7.33(4H,m)[α]_(D) ²⁰: −32.8 (c=0.19, MeOH)

EXAMPLE 402-((2,6-dichlorophenyl)carbonylamino)-5-oxo-5-(2,4,8-triaza-1-oxo-4-phenylspiro[4.5]dec-8-yl)pentanoicacid (40)

In 2 ml of dichloromethane, 436 mg of t-butyl2-((2,6-dichlorophenyl)carbonylamino)-5-oxo-5-(2,4,8-triaza-1-oxo-4-phenylspiro[4.5]dec-8-yl)pentanoicacid was dissolved, and 1.5 ml of trifluoroacetic acid was addedthereto, followed by stirring the resulting mixture at room temperaturefor 4 hours. After concentrating the reaction solution, the residue wasreprecipitated from dichloromethane/ether to obtain 295 mg of2-((2,6-dichlorophenyl)carbonylamino)-5-oxo-5-(2,4,8-triaza-1-oxo-4-phenylspiro[4.5]dec-8-yl)pentanoicacid (yield: 75%).

LR-MS(m/z): 531 (M⁺−H) IR(KBr): 3258, 3066, 2929, 1716, 1657, 1625,1601, 1501, 1433, 1381, 1366, 1194, 1089, 801, 751 cm⁻¹ NMR(300MHz,CDCl₃, δ ppm): NMR(300 MHz,CDCl₃, δ ppm):1.71-1.85(2H,m),2.00-2.22(1H,m),2.35-2.72(4H,m),2.78-2.83(1H,m),3.40-3.48(1H,m),3.78-4.00(2H,m),4.43-4.58(1H,m),4.70-4.82(1H,m),4.73(2H,s),6.72-6.93(3H,m),7.20-7.35(5H,m)HR-MS: C₂₅H₂₅Cl₂N₄O₅ Calcd.: 531.1202. Found: 531.1218. [α]_(D) ²⁰:−53.6° (c=0.04, MeOH)

EXAMPLE 41 Methyl2-((2,6-dichlorophenyl)carbonylamino)-3-((2,4,8-triaza-1-oxo-4-phenyl-2-prop-2-enylspiro[4.5]dec-8-yl)carbonylamino)propionate(41)

In 3 ml of dichloromethane and 2 ml of acetonitrile, 67 mg of methyl3-amino-2-((2,6-dichlorophenyl)carbonylamino)propionate was dissolved,and 31 mg of saturated sodium hydrogen carbonate and 56 mg ofchloroformic acid p-nitrophenyl ester were added thereto under argonatmosphere, followed by stirring the resulting mixture at roomtemperature for 2.5 hours. To the reaction mixture, 95 mg of2,4,8-triaza-4-phenyl-2-prop-2-enylspiro[4.5]decane-1-one and 167 μl oftriethylamine were added, and the resulting mixture was stirredovernight at room temperature. After concentrating the reaction mixture,saturated aqueous sodium hydrogen carbonate solution was added, and theresulting mixture was extracted with chloroform. Organic layers werecombined, washed with 0.1N hydrochloric acid and with saturated saline,dried over anhydrous sodium sulfate, and concentrated. The residue waspurified by column chromatography (dichloromethane/methanol=60:1) toobtain 89.8 mg of methyl2-((2,6-dichlorophenyl)carbonylamino)-3-((2,4,8-triaza-1-oxo-4-phenyl-2-prop-2-enylspiro[4.5]dec-8-yl)carbonylamino)propionate(yield: 66%).

LR-MS(m/z): 587(M⁺) IR(KBr): 3384, 3065, 2927, 1746, 1696, 1602, 1531,1470, 1432, 1378, 1260, 1198, 1171, 1128, 802, 749 cm⁻¹ NMR(300MHz,CDCl₃, δ ppm): 1.63-1.73(2H,m),2.45-2.60(2H,m),3.60-3.92(6H,m),3.82(3H,s),4.02-4.07(2H,m),4.65(2H,s),4.80-4.87(1H,m),5.24-5.32(2H,m),5.38(1H,brs),5.75-5.86(1H,m),6.73-6.80(2H,m),6.82-6.90(1H,m),7.22-7.34(5H,m),7.45(1H,brs) [α]_(D) ²⁰: −8.03° (c=0.07, MeOH)

EXAMPLE 422-((2,6-dichlorophenyl)carbonylamino)-3-((2,4,8-triaza-1-oxo-4-phenyl-2-prop-2-enylspiro[4.5]dec-8-yl)carbonylamino)propanoicacid (42)

In 3 ml of methanol, 86.3 mg of methyl2-((2,6-dichlorophenyl)carbonylamino)-3-((2,4,8-triaza-1-oxo-4-phenyl-2-prop-2-enylspiro[4.5]dec-8-yl)carbonylamino)propionatewas dissolved, and 0.75 ml of 1N aqueous sodium hydroxide solution wasadded thereto, followed by stirring the resulting mixture at roomtemperature for 1 hour. To the reaction solution, 1N hydrochloric acidwas added, and the resulting mixture was extracted with chloroform.Organic phases were combined, washed with saturated saline, dried overanhydrous sodium sulfate, and concentrated. The residue wasreprecipitated from chloroform/n-hexane to obtain 104 mg of2-((2,6-dichlorophenyl)carbonylamino)-3-((2,4,8-triaza-1-oxo-4-phenyl-2-prop-2-enylspiro[4.5]dec-8-yl)carbonylamino)propanoicacid (yield: %).

LR-MS(m/z): 572(M⁺−H) IR(KBr): 3384, 2928, 1686, 1601, 1539, 1473, 1432,1379, 1262, 1197, 801,749 cm⁻¹ NMR(300 MHz,CDCl₃, δ ppm):1.69-1.82(2H,m), 2.44-2.62(2H,m), 3.48-3.57(1H,m), 3.71-4.01(5H,m),4.02-4.05(2H,m), 4.51-4.58(1H,m), 4.67(2H,s), 5.25-5.36)2H,m),5.77-5.90(1H,m), 5.95(1H,brs), 6.73-6.81(2H,m), 6.54-6.97(1H,m),7.23-7.37(5H,m), 7.63(1H,brs) HR-MS: C₂₇H₂₉Cl₂N₅O₅ Calcd.: 572.1467.Found: 572.1417. [α]_(D) ²⁰: −55.8° (c=0.04, MeOH)

EXAMPLE 432-(((S)-3-acetylthiazolidin-4-carbonyl)amino)-3-((3-methyl-4-oxo-1-phenyl-1,3,8-triazaspiro[4.5]decane-8-carbonyl)amino)propionicacid methyl ester (43)

To a solution of 331 mg (0.85 mmol) of2-amino-3-((3-methyl-4-oxo-1-phenyl-1,3,8-triazaspiro[4.5]decane-8-carbonyl)amino)propionicacid methyl ester in 8.5 ml of dichloromethane, 149 mg (0.85 mmol) of(S)-3-acetylthiazolidin-4-carboxylic acid, 376 mg (0.85 mmol) of BOPreagent and 0.16 ml (0.95 mmol) of N,N-diisopropylethylamine were added,and the resulting mixture was stirred overnight at room temperature. Tothe reaction mixture, 10 ml of semi-saturated saline was added, and theresulting mixture was extracted with ethyl acetate. Organic phases werecombined, washed with 0.5M hydrochloric acid, with saturated aqueoussodium hydrogen carbonate solution and with saturated saline, dried overanhydrous sodium sulfate, and concentrated. The residue was purified bysilica gel column chromatography (chloroform:methanol=100:1-10:1) toobtain 305 mg of2-(((S)-3-acetylthiazolidin-4-carbonyl)amino)-3-((3-methyl-4-oxo-1-phenyl-1,3,8-triazaspiro[4.5]decane-8-carbonyl)amino)propionicacid methyl ester (yield: 65%).

LR-MS (m/z): 547 (M+H)⁺IR(KBr): 3376, 1750, 1699, 1651, 1639, 1532,1404, 1268, 1204, 1153, 1128, 1055, 986 cm⁻¹ NMR (300 MHz, CDCl₃, δppm): 8.07 (1H, d, J=6.2), 7.50-7.12 (3H, m), 6.87 (1H, t, J=7.1 Hz),6.76 (2H, d, J=8.2), 5.15 (1H, t, J=5.5), 5.00 (1H, dd, J=7.0, 2.9),4.73-4.42 (5H, m), 3.93-3.15 (12H, m), 3.02 (3H, s), 2.66-2.46 (2H, m),2.17 (3H,s)

EXAMPLE 442-(((S)-3-acetylthiazolidine-4-carbonyl)amino)-3-((3-methyl-4-oxo-1-phenyl-1,3,8-triazaspiro[4.5]decane-8-carbonyl)amino)propionicacid (44)

To a solution of 176 mg (0.32 mmol) of2-(((S)-3-acetylthiazolidin-4-carbonyl)amino)-3-((3-methyl-4-oxo-1-phenyl-1,3,8-triazaspiro[4.5]decane-8-carbonyl)amino)propionicacid methyl ester in 4 ml of tetrahydrofuran, 4 ml of 0.1M aqueouslithium hydroxide solution was added, and the resulting mixture wasstirred overnight at room temperature. To the reaction mixture, 0.1Mhydrochloric acid was added, and the resulting mixture was extractedwith ethyl acetate. Organic phases were combined, washed with saturatedsaline, dried over anhydrous sodium sulfate, and concentrated. Theresidue was reprecipitated from ether/chloroform to obtain 130 mg of2-(3-acetylthiazolidine-4-carbonyl)amino)-3-((3-methyl-4-oxo-1-phenyl-1,3,8-triazaspiro[4.5]decane-8-carbonyl)amino)propionicacid (yield: 76%).

LR-MS (m/z): 533 (M+H)⁺IR (KBr): 3374, 1689, 1633, 1533, 1407, 1268,1198, 1155, 1087, 1056, 987 cm⁻¹ NMR (300 MHz, CDCl₃, δppm): 8.79 (1H,d, J=4.7), 7.48-7.13 (2H, m), 6.86 (1H, t, J=7.2), 6.73 (2H, d, J=7.2),5.81 (1H, t, J6.2), 4.99 (1H, dd, J=6.8, 3.0), 4.68 (2H, s), 4.61 (1H,d, J=8.5), 4.54 (1H, d,J=8.5), 4.27 (1H, m), 3.95-3.54 (6H, m),3.38-3.16 (2H, m), 3.01 (3H, s), 2.62-2.42 (2H, m), 2.21 (3H, s), 1.69(2H, d, J=14.6)

EXAMPLE 452-(((R)-3-acetylthiazolidin-4-carbonyl)amino)-3-((3-methyl-4-oxo-1-phenyl-1,3,8-triazaspiro[4.5]decane-8-carbonyl)amino)propionicacid methyl ester (45)

To a solution of 301 mg (0.77 mmol) of2-amino-3-((3-methyl-4-oxo-1-phenyl-1,3,8-triazaspiro[4.5]decane-8-carbonyl)amino)propionicacid methyl ester in 7.5 ml of dichloromethane, 135 mg (0.77 mmol) of(R)-3-acetylthiazolidin-4-carboxylic acid, 342 mg (0.77 mmol) of BOPreagent and 0.15 ml (0.86 mmol) of N,N-diisopropylethylamine were added,and the resulting mixture was stirred overnight at room temperature. Tothe reaction mixture, 10 ml of semi-saturated saline was added, and theresulting mixture was extracted with ethyl acetate. Organic phases werecombined, washed with 0.5M hydrochloric acid, with saturated aqueoussodium hydrogen carbonate solution and with saturated saline, dried overanhydrous sodium sulfate, and concentrated. The residue was purified bysilica gel column chromatography (chloroform:methanol=100:1-10:1) toobtain 309 mg of2-(((R)-3-acetylthiazolidin-4-carbonyl)amino)-3-((3-methyl-4-oxo-1-phenyl-1,3,8-triazaspiro[4.5]decane-8-carbonyl)amino)propionicacid methyl ester (yield: 73%).

LR-MS (m/z): 547 (M+H)⁺IR (KBr): 3372, 1749, 1698, 1658, 1634, 1531,1438, 1404, 1266, 1207, 1153, 1128, 1084, 1055, 986 cm⁻¹ NMR (300 MHz,CDCl₃, δ ppm): 8.98 (0.33H, d, J=5.3), 7.63 (0.67H, d, J=7.0), 7.48-7.13(3H, m), 6.91-6.72 (3H, m), 5.45 (0.67H, t, J=6.2), 4.98-4.48 (6.33H,m), 4.00-3.32 (11H, m), 3.16 (1H, dd, J=11.5, 6.5 Hz), 3.01 (3H, s),2.65-2.42 (2H, m), 2.18 (1H, s), 2.15 (2H, s)

EXAMPLE 46 Methyl2-(((R)-3-acetylthiazolidin-4-carbonyl)amino)-3-((3-methyl-4-oxo-1-phenyl-1,3,8-triazaspiro[4.5]decane-8-carbonyl)amino)propionate(46)

To a solution of 171 mg (0.31 mmol) of2-(3-acetylthiazolidin-4-carbonyl)amino)-3-((3-methyl-4-oxo-1-phenyl-1,3,8-triazaspiro[4.5]decane-8-carbonyl)amino)propionicacid methyl ester in 4 ml of tetrahydrofuran, 4 ml of 0.1M aqueouslithium hydroxide solution was added, and the resulting mixture wasstirred overnight at room temperature. To the reaction mixture, 0.1Mhydrochloric acid was added, and the resulting mixture was extractedwith ethyl acetate. Organic phases were combined, washed with saturatedsaline, dried over anhydrous sodium sulfate, and concentrated. Theresidue was reprecipitated from ether/chloroform to obtain 135 mg of2-(3-acetylthiazolidin-4-carbonyl)amino)-3-((3-methyl-4-oxo-1-phenyl-1,3,8-triazaspiro[4.5]decane-8-carbonyl)amino)propionicacid (yield: 81%).

LR-MS (m/z): 533 (M+H)⁺IR (KBr): 3367, 1693, 1633, 1532, 1408, 1267,1204, 1154, 1124, 1056, 986 cm⁻¹ NMR (300 MHz, CDCl₃, δppm): 9.17(0.25H, d, J=5.0), 8.38 (0.75H, d, J=5.0), 7.48-7.13 (2H, m), 6.87 (1H,t, J=7.3), 6.74 (2H, d, J=8.2), 5.82 (1H, m), 4.92-4.33 (6H, m),3.88-3.54 (6H, m), 3.31 (1H, dd, J=11.9, 4.0), 3.20 (1H, dd, J=11.9,6.9), 3.01 (3H, s), 2.62-2.42 (2H, m), 2.17 (3H, s), 1.68 (2H, d,J=13.2)

EXAMPLE 472-benzyloxycarbonylamino-3-((3-methyl-4-oxo-1-phenyl-1,3,8,-triazaspiro[4.5]decane-8-carbonyl)amino)propionicacid methyl ester (47)

To a solution of 288 mg (0.74 mmol) of2-amino-3-((3-methyl-4-oxo-1-phenyl-1,3,8-triazaspiro[4.5]decane-8-carbonyl)amino)propionicacid methyl ester in 4 ml of dichloromethane, 0.21 ml (1.48 mmol) oftriethylamine and 0.1 ml (0.89 mmol) of benzyloxycarbonyl chloride wereadded, and the resulting mixture was stirred at room temperature for oneday. To the reaction mixture, saturated aqueous sodium hydrogencarbonate solution was added, and the resulting mixture was extractedwith chloroform. Organic phases were combined, washed with 10% aqueouscitric acid solution and with saturated saline, dried over anhydroussodium sulfate, and concentrated. The residue was purified by silica gelcolumn chromatography (chloroform:methanol=60:1) to obtain 284 mg of2-benzyloxycarbonylamino-3-((3-methyl-4-oxo-1-phenyl-1,3,8,-triazaspiro[4.5]decane-8-carbonyl)amino)propionicacid methyl ester (yield: 73%)

LR-MS (m/z): 524 (M+H)⁺NMR (300 MHz, CDCl₃, δ ppm): 7.40-7.22 (7H, m),6.94 (1H, d, J=6.9), 6.74 (2H, d, J=8.2), 6.31 (1H, br d, J=6.6), 5.10(1H, br s), 5.09 (2H, s), 3.92-3.57 (9H, m), 3.01 (3H, s), 2.62-2.48(2H, m), 1.66 (2H, d, J=13.5)

EXAMPLE 482-benzyloxycarbonylamino-3-((3-methyl-4-oxo-1-phenyl-1,3,8,-triazaspiro[4.5]decane-8-carbonyl)amino)propionicacid (48)

To a solution of 177 mg (0.34 mmol) of2-benzyloxycarbonylamino-3-((3-methyl-4-oxo-1-phenyl-1,3,8,-triazaspiro[4.5]decane-8-carbonyl)amino)propionicacid methyl ester in 4 ml of tetrahydrofuran, 4 ml of 0.1M aqueouslithium hydroxide solution was added, and the resulting mixture wasstirred overnight at 0° C. To the reaction mixture, 0.1M hydrochloricacid was added, and the resulting mixture was extracted with ethylacetate. Organic phases were combined, washed with saturated saline,dried over anhydrous sodium sulfate, and concentrated. The residue wasreprecipitated from ether/chloroform to obtain 143 mg of2-benzyloxycarbonylamino-3-((3-methyl-4-oxo-1-phenyl-1,3,8,-triazaspiro[4.5]decane-8-carbonyl)amino)propionicacid (yield: 83%).

LR-MS (m/z): 510 (M+H)⁺NMR (300 MHz, CDCl₃, δppm): 7.38-7.20 (7H, m),6.86 (1H, t, J=7.4), 6.73 (2H, d, J=8.2), 6.54 (1H, d, J=5.0), 5.66 (1H,br s), 5.09 (2H, s), 4.66 (2H, s), 4.24 (1H, br s), 3.90-3.60 (5H, m),3.45 (1H, br d, J=12.3), 3.00 (3H, s), 2.60-2.40 (2H, m), 1.76-1.56 (2H,m)

EXAMPLE 492-(2,6-dichlorobenzylamino)-3-((3-methyl-4-oxo-1-phenyl-1,3,8-triazaspiro[4.5]decane-8-carbonyl)amino)propionicacid methyl ester (49)

To a solution of 288 mg (0.74 mmol) of2-amino-3-((3-methyl-4-oxo-1-phenyl-1,3,8-triazaspiro[4.5]decane-8-carbonyl)amino)propionicacid methyl ester in 4 ml of dichloromethane, 0.21 ml (1.48 mmol) oftriethylamine and 214 mg (0.89 mmol) of 2,6-dichlorobenzyl bromide wereadded, and the resulting mixture was stirred at room temperature for oneday. To the reaction mixture, saturated aqueous sodium hydrogencarbonate solution was added, and the resulting mixture was extractedwith chloroform. Organic phases were combined, washed with saturatedsaline, dried over anhydrous sodium sulfate, and concentrated. Theresidue was purified by silica gel column chromatography(chloroform:methanol=60:1) to obtain 195 mg of2-(2,6-dichlorobenzylamino)-3-((3-methyl-4-oxo-1-phenyl-1,3,8-triazaspiro[4.5]decane-8-carbonyl)amino)propionicacid methyl ester (yield: 48%).

LR-MS (m/z): 548 (M+H)⁺NMR (300 MHz, CDCl₃, δppm): 7.31-7.23 (4H, m),7.11 (1H, dd, J=8.8, 7.3), 6.85 (1H, t, J=7.5 Hz), 6.73 (2H, d, J=7.9),5.28 (1H, dd, J=6.1, 3.8), 4.68 (2H, s), 4.17 (1H, d, J=12.6), 3.99 (1H,d, J=12.6), 3.94-3.58 (5H, m), 3.72 (3H, s), 3.48 (1H, dd, J=7.6, 4.4),3.26 (1H, ddd, J=13.2, 7.9, 3.8), 3.01 (3H, s), 2.60-2.46 (2H, m), 1.65(2H, d, J=13.8)

EXAMPLE 502-(2,6-dichlorobenzylamino)-3-((3-methyl-4-oxo-1-phenyl-1,3,8-triazaspiro[4.5]decane-8-carbonyl)amino)propionicacid (50)

To a solution of 132 mg (0.24 mmol) of2-(2,6-dichlorobenzylamino)-3-((3-methyl-4-oxo-1-phenyl-1,3,8-triazaspiro[4.5]decane-8-carbonyl)amino)propionicacid methyl ester in 3 ml of tetrahydrofuran, 2.6 ml of 0.1M aqueouslithium hydroxide solution was added, and the resulting mixture wasstirred overnight at room temperature. To the reaction mixture, 0.1Mhydrochloric acid was added, and the resulting mixture was extractedwith chloroform. Organic phases were combined, washed with saturatedsaline, dried over anhydrous sodium sulfate, and concentrated. Theresidue was reprecipitated from ether/chloroform to obtain 103 mg of2-(2,6-dichlorobenzylamino)-3-((3-methyl-4-oxo-1-phenyl-1,3,8-triazaspiro[4.5]decane-8-carbonyl)amino)propionicacid (yield: 81%).

LR-MS (m/z): 534 (M+H)⁺NMR (300 MHz, CDCl₃, δppm): 7.40-7.24 (5H, m),6.85 (1H, t, J=7.3), 6.73 (2H, d, J=7.9), 6.71 (1H, br s), 4.65 (2H, s),4.47 (1H, d, J=13.5), 4.41 (1H, d, J=13.5), 4.01-3.56 (7H, m), 3.00 (3H,s), 2.56-2.40 (2H, m), 1.67 (2H, d, J=14.9)

EXAMPLE 51 Inhibitory Effect by Compounds Against Binding between CS-1Peptide and VLA-4-IgG Chimera Protein

In accordance with the teaching of a report (Humphrise, M. J. et al.J.Bio.Chem.,262,6886-6892(1987)), a conjugate between a peptide (Gys LeuHis Gly Pro Glu Glu Ile Leu Asp Val Pro Ser Thr) containing CS-1sequence and rabbit IgG (Sigma) was prepared. This was diluted withphosphate buffer (hereinafter referred to as “PBS(−)” for short), andthe obtained solution was placed in the wells of a 96-well immunoplate(NUNC) in an amount of 100 μl/well, followed by leaving to stand theimmunoplate at 4° C. for 16 hours to immobilize the conjugate.

The wells were then washed twice with PBS(−), and 1% BSA solution inPBS, which BSA was heated at 80° C. for 10 minutes, was placed in eachwell in an amount of 300 μl/well. The immunoplate was left to stand at4° C. for 3 hours, and then the solution in each well was removed bysuction.

Each compound and VLA-4-IgG chimera protein (100 μl) were preliminarilyreacted at room temperature for 20 minutes, and then the resultingmixture was allowed to react with the CS-1peptide in each well at 30° C.for 3 hours. Thereafter, non-bound VLA-4-IgG chimera protein was removedby suction, and each well was washed twice with 0.1% BSA-containing TBSbuffer (150 mM NaCl, 25 mM Tris-HCl, 1 mM MnCl₂, PH7.4). To the boundVLA-4-IgG chimera protein, biotin-labelled anti-human IgG antibody(Vector) as a primary antibody was added, and then avidin-labelled horseradish peroxidase (Sigma) as a secondary antibody was added, therebyallowing the reactions. Then o-phenylenediamine as a substrate was addedto color the reaction solution, and the absorbance at 490 nm wasmeasured. From this absorbance, the binding inhibitory activity of eachcompound was determined. The inhibitory activities of the representativecompounds are shown in Table 1.

TABLE 1 Compound No. Inhibitory Activity (IC₅₀: nM) 3 6.1 7 18 13 2.9 272.1 31 0.98 Industrial Field

The novel spiro derivatives according to the present invention haveactivities to inhibit cell adhesion via adhesion molecules, especiallyadhesion molecule VLA-4. Since the spiro acid derivatives according tothe present invention are excellent in the effect of inhibiting celladhesion via adhesion molecules, they are useful as therapeutic drugsagainst various inflammatory diseases.

1. A spiro derivative represented by the Formula I:

(wherein 1 and m independently represent integers of 0 to 2; nrepresents; A represents an oxygen atom, —CH—, carbon having a doublebond or a nitrogen atom (with the proviso that when A is an oxygen atom,R₃ does not exist); B represents —CH₂— or —NH—; C′ and D representhydrogen, or C′ and D cooperatively represent O; X₁ and Y₁ independentlyrepresent hydrogen, halogen, C₁-C₈ alkyl, C₁-C₈ alkoxy, cyano, nitro,hydroxyl, amino or tetrazole; R₁ and R₂ independently represent hydrogenor C₁-C₆ linear alkyl; R₃ and R₄ independently represent hydrogen, C₁-C₆linear alkyl, C₃-C₈ branched alkyl, or phenyl or benzyl, this phenyl orbenzyl being substituted with 0 to 2 substituents selected from thegroup consisting of halogen, C₁-C₈ alkyl, C₁-C₈ alkoxy, cyano, nitro,hydroxyl, amino and tetrazole; F represents —CH₂— or —C(O)—; when A is anitrogen atom, R₃, A and R4 may cooperatively represent (i) Formula II:

(wherein p represents an integer of 0 to 4; E represents a carbon atomor a nitrogen atom; R₆ and R₇ independently represent hydrogen, C₁-C₆linear alkyl, C₃-C₈ branched alkyl, C₁-C₆ linear acyl, C₃-C₈ branchedacyl, pyrrolidinecarbonyl, piperidinecarbonyl, or phenyl,phenylsulfonyl, benzoyl, benzyl, indole or N-phenylamide, this phenyl,phenylsulfonyl, benzoyl, benzyl, indole or N-phenylamide beingsubstituted with 0 to 2 substituents selected from the group consistingof halogen, C₁-C₈ alkyl, C₁-C₈ alkoxy, cyano, nitro, hydroxyl, amino andtetrazole, or Formula III:

(wherein R₈ represents hydrogen, C₁-C₆ linear alkyl, C₃-C₈ branchedalkyl, or phenyl or benzyl, this phenyl or benzyl being substituted with0 to 2 substituents selected from the group consisting of halogen,halogen, C₁-C₈ alkyl, C₁-C₈ alkoxy, cyano, nitro, hydroxyl, amino andtetrazole), (ii) Formula IV:

(wherein R₉ represents C₁-C₆ linear alkyl, C₃-C₈ branched alkyl, C₁-C₆linear acyl, C₃-C₈ branched acyl, C₅-C₇ cycloalkylcarbonyl, C₁-C₆ linearalkylsulfonyl, C₃-C₈ branched alkylsulfonyl, or benzoyl, phenylsulfonylor benzyl, this benzoyl, phenylsulfonyl or benzyl being substituted with0 to 2 substituents selected from the group consisting of halogen, C₁-C₈alkyl, C₁-C₈ alkoxy, cyano, nitro, hydroxyl, amino and tetrazole) (iii)Formula V:

(wherein R₁₀ represents hydrogen, C₁-C₆ linear alkyl, C₃-C₈ branchedalkyl, C₆-C₁₀ alkylcycloalkyl, or phenyl or benzyl, this phenyl orbenzyl being substituted with 0 to 2 substituents selected from thegroup consisting of halogen, C₁-C₈ alkyl, C₁-C₈ alkoxy, cyano, nitro,hydroxyl, amino and tetrazole); When A is —CH— or carbon having a doublebond, R₃, A and R₄ may cooperatively form adamantyl, or Formula VI:

(wherein definitions of X₂ and Y₂ are the same as those of X₁ and Y₁,respectively)

 or Formula VII: (wherein definition of R₁₁ is the same as that of R₉);R₅ represents hydrogen, C₁-C₆ linear alkyl, C₃-C₈ branched alkyl, allyl,homoallyl, C₆-C₁₀ alkylcycloalkyl, or phenyl, benzyl, phenethyl, styrylor naphthylmethyl, this phenyl, benzyl, phenethyl, styryl ornaphthylmethyl being substituted with 0 to 2 substituents selected fromthe group consisting of halogen, C₁-C₈ alkyl, C₁-C₈ alkoxy, cyano,nitro, hydroxyl, amino and or a pharmaceutically acceptable saltthereof.
 2. The spiro derivative or the pharmaceutically acceptable saltthereof according to claim 1, wherein F represents —C(O)—; A represents—CH—, carbon having a double bond or nitrogen atom, B represents —CH₂—or —NH—; C′ and D represent hydrogen or C′ and D cooperatively representO; X and Y independently represent hydrogen, halogen, methyl, methoxy,cyano, nitro, hydroxyl, amino or tetrazole; R₁ and R₂ independentlyrepresent hydrogen or C₁-C₆ linear alkyl; R₃ and R₄ independentlyrepresent hydrogen, C₁-C₆ linear alkyl or C₃-C₈ branched alkyl, orphenyl or benzyl, this phenyl or benzyl being substituted with 0 to 2substituents selected from the group consisting of halogen, methyl,methoxy, cyano, nitro, hydroxyl, amino and tetrazole; when A is anitrogen atom, R₃, A and R₄ may cooperatively represent (i) said FormulaII (wherein p represents an integer of 0 to 4; E represents carbon atomor nitrogen atom; R₆ and R₇ independently represent hydrogen, C₁-C₆linear alkyl, C₃-C₈ branched alkyl, C₁-C₆ linear acyl, C₃-C₈ branchedacyl, pyrrolidinecarbonyl, piperidinecarbonyl, or phenyl,phenylsulfonyl, benzoyl, benzyl, indole or N-phenylamide, this phenyl,phenylsulfonyl, benzoyl, benzyl, indole or N-phenylamide beingsubstituted with 0 to 2 substituents selected from the group consistingof halogen, methyl, methoxy, cyano, nitro, hydroxyl, amino andtetrazole, or said Formula III (wherein R₈ represents hydrogen, C₁-C₆linear alkyl, C₃-C₈ branched alkyl, or phenyl or benzyl, this phenyl orbenzyl being substituted with 0 to 2 substituents selected from thegroup consisting of halogen, methyl, methoxy, cyano, nitro, hydroxyl,amino and tetrazole), (ii) said Formula IV (wherein R₉ represents C₁-C₆linear alkyl, C₃-C₈ branched alkyl, C₁-C₆ linear acyl, C₃-C₈ branchedacyl, C₅-C₇ cycloalkylcarbonyl, C₁-C₆ linear alkylsulfonyl, C₃-C₈branched alkylsulfonyl, or benzoyl, phenylsulfonyl or benzyl, thisbenzoyl, phenylsulfonyl or benzyl being substituted with 0 to 2substituents selected from the group consisting of halogen, methyl,methoxy, cyano, nitro, hydroxyl, amino and tetrazole), (iii) saidFormula V: (wherein R₁₀ represents hydrogen, C₁-C₆ linear alkyl, C₃-C₈branched alkyl, C₆-C₁₀ alkylcycloalkyl, or phenyl or benzyl, this phenylor benzyl being substituted with 0 to 2 substituents selected from thegroup consisting of halogen, methyl, methoxy, cyano, nitro, hydroxyl,amino and tetrazole); when A is —CH— or carbon having a double bond, R₃,A and R₄ may cooperatively represent adamantyl or said Formula VI(wherein X₂ and Y₂ represent the same definitions as described above);R₅ represents hydrogen, C₁-C₆ linear alkyl, C₃-C₈ branched alkyl, allyl,homoallyl, C₆-C₁₀ alkylcycloalkyl, or phenyl, benzyl, phenethyl, styrylor naphthylmethyl, this phenyl, benzyl, phenethyl, styryl ornaphthylmethyl being substituted with 0 to 2 substituents selected fromthe group consisting of halogen, methyl, methoxy, cyano, nitro,hydroxyl, amino and tetrazole.
 3. The spiro derivative or thepharmaceutically acceptable salt thereof according to claim 1 or 2,wherein A represents a nitrogen atom, R₃, A and R₄ cooperativelyrepresent (i) said Formula II (wherein R₆ and R₇ independently representhydrogen, C₁-C₃ linear alkyl, C₃-C₆ branched alkyl, C₁-C₃ linear acyl,C₃-C₆ branched acyl, pyrrolidinecarbonyl, piperidinecarbonyl, or phenyl,benzoyl, benzyl, indole or N-phenylamide, this phenyl, benzoyl, benzyl,indole or N-phenylamide being substituted with 0 to 2 substituentsselected from the group consisting of halogen, methyl, methoxy, cyano,nitro, hydroxyl, amino and tetrazole), or said Formula III (wherein R₈represents hydrogen, C₁-C₃ linear alkyl, C₃-C₆ branched alkyl, or phenylor benzyl, this phenyl or benzyl being substituted with 0 to 2substituents selected from the group consisting of halogen, methyl,methoxy, cyanc, nitro, hydroxyl, amino and tetrazole), (ii) said FormulaIV (wherein R₉ represents C₁-C₃ linear alkyl, C₃-C₆ branched alkyl,C₁-C₃ linear acyl, C₃-C₆ branched acyl, C₅-C₇ cycloalkylcarbonyl, C₁-C₃linear alkylsulfonyl, C₃-C₈ branched alkylsulfonyl, or benzoyl orbenzyl, this benzoyl or benzyl being substituted with 0 to 2substituents selected from the group consisting of halogen, methyl,methoxy, cyano, nitro, hydroxyl, amino and tetrazole), or (iii) saidFormula V (wherein R₁₀ represents hydrogen, C₁-C₃ linear alkyl, C₃-C₆branched alkyl, C₆-C₁₀ alkylcycloalkyl, or phenyl or benzyl, this phenylor benzyl being substituted with 0 to 2 substituents selected from thegroup consisting of halogen, methyl, methoxy, cyano, nitro, hydroxyl,amino and tetrazole), the definitions of the symbols other thanmentioned above are the same as those described in claim 1 or
 2. 4. Thespiro derivative or the pharmaceutically acceptable salt thereofaccording to claim 1 or 2, wherein A represents a nitrogen atom, R₃, Aand R₄ cooperatively represent (i) said Formula II (wherein R₆ and R₇independently represent hydrogen, C₁-C₃ linear acyl, C₃-C₆ branchedacyl, pyrrolidinecarbonyl, piperidinecarbonyl, or phenyl, benzoyl,benzyl, indole or N-phenylamide, this phenyl, benzoyl, benzyl, indole orN-phenylamide being substituted with 0 to 2 substituents selected fromthe group consisting of halogen, methyl, methoxy, cyano, nitro,hydroxyl, amino and tetrazole), or said Formula III (wherein R₈represents hydrogen, C₁-C₃ linear alkyl, C₃-C₆ branched alkyl, or benzylsubstituted with 0 to 2 substituents selected from the group consistingof halogen, methyl, methoxy, cyano, nitro, hydroxyl, amino andtetrazole), (ii) said Formula IV (wherein R₉ represents C₁-C₃ linearacyl, C₃-C₆ branched acyl, C₅-C₇ cycloalkylcarbonyl, or benzoyl orbenzyl, this benzoyl or benzyl being substituted with 0 to 2substituents selected from the group consisting of halogen, methyl,methoxy, cyano, nitro, hydroxyl, amino and tetrazole), or (iii) saidFormula V (wherein R₁₀ represents hydrogen, C₃-C₃ linear alkyl, C₃-C₆branched alkyl, C₆-C₁₀ alkylcycloalkyl, or phenyl or benzyl, this phenylor benzyl being substituted with 0 to 2 substituents selected from thegroup consisting of halogen, methyl, methoxy, cyano, nitro, hydroxyl,amino and tetrazole), the definitions of the symbols other thanmentioned above are the same as those described in claim 1 or
 2. 5. Apharmaceutical comprising said spiro derivative or a pharmaceuticallyacceptable salt thereof according to claim 1 or 2 as an effectiveingredient and a carrier.
 6. A method for inhibiting an VLA-4 adhesionmolecule, comprising administering an effective amount of a spiroderivative or a pharmaceutically acceptable salt thereof, wherein thespiro derivative is represented by Formula I:

(wherein 1 and m independently represent integers of 0 to 2; nrepresents an integer of 1 to 3; A represents an oxygen atom, —CH—,carbon having a double bond or a nitrogen atom (with the proviso thatwhen A is an oxygen atom, R3 does not exist); B represents —CH₂— or—NH—; C′ and D represent hydrogen, or C′ and D cooperatively representO; X₁ and Y₁ independently represent hydrogen, halogen, C₁-C₈ alkyl,C₁-C₈ alkoxy, cyano, nitro, hydroxyl, amino or tetrazole; R₁ and R₂independently represent hydrogen or C₁-C₆ linear alkyl; R₃ and R₄independently represent hydrogen, C₁-C₆ linear alkyl, C₃-C₈ branchedalkyl, or phenyl or benzyl, this phenyl or benzyl being substituted with0 to 2 substituents selected from the group consisting of halogen, C₁-C₈alkyl, C₁-C₈ alkoxy, cyano, nitro, hydroxyl, amino and tetrazole; Frepresents —CH₂— or —C(O)—; when A is a nitrogen atom, R₃, A and R₄ maycooperatively represent (i) Formula II:

(wherein p represents an integer of 0 to 4; E represents a carbon atomor a nitrogen atom; R₆ and R₇ independently represent hydrogen, C₁-C₆linear alkyl, C₃-C₈ branched alkyl, C₁-C₆ linear acyl, C₃-C₈ branchedacyl, pyrrolidinecarbonyl, piperidinecarbonyl, or phenyl,phenylsulfonyl, benzoyl, benzyl, indole or N-phenylamide, this phenyl,phenylsulfonyl, benzoyl, benzyl, indole or N-phenylamide beingsubstituted with 0 to 2 substituents selected from the group consistingof halogen, C₁-C₈ alkyl, C₁-C₈ alkoxy, cyano, nitro, hydroxyl, amino andtetrazole, or Formula III:

(wherein R₈ represents hydrogen, C₁-C₆ linear alkyl, C₃-C₈ branchedalkyl, or phenyl or benzyl, this phenyl or benzyl being substituted with0 to 2 substituents selected from the group consisting of halogen,halogen, C₁-C₈ alkyl, C₁-C₈ alkoxy, cyano, nitro, hydroxyl, amino andtetrazole), (ii) Formula IV:

(wherein R₉ represents C₁-C₆ linear alkyl, C₃-C₈ branched alkyl, C₁-C₆linear acyl, C₃-C₈ branched acyl, C₅-C₇ cycloalkylcarbonyl, C₁-C₆ linearalkylsulfonyl, C₃-C₈ branched alkylsulfonyl, or benzoyl, phenylsulfonylor benzyl, this benzoyl, phenylsulfonyl or benzyl being substituted with0 to 2 substituents selected from the group consisting of halogen, C₁-C₈alkyl, C₁-C₈ alkoxy, cyano, nitro, hydroxyl, amino and tetrazole) (iii)Formula V:

(wherein R₁₀ represents hydrogen, C₁-C₆ linear alkyl, C₃-C₈ branchedalkyl, C₆-C₁₀ alkylcycloalkyl, or phenyl or benzyl, this phenyl orbenzyl being substituted with 0 to 2 substituents selected from thegroup consisting of halogen, C₁-C₈ alkyl, C₁-C₈ alkoxy, cyano, nitro,hydroxyl, amino and tetrazole); When A is —CH— or carbon having a doublebond, R₃, A and R₄ may cooperatively form adamantyl, or Formula VI:

(wherein definitions of X₂ and Y₂ are the same as those of X₁ and Y₁,respectively)

 or Formula VII: (wherein definition of R₁₁ is the same as that at R₉);R₅ represents hydrogen, C₁-C₆ linear alkyl, C₃-C₈ branched alkyl, allyl,homoallyl, C₆-C₁₀ alkylcycloalkyl, or phenyl, benzyl, phenethyl, styrylor naphthylmethyl, this phenyl, benzyl, phenethyl, styryl ornaphthylmethyl being substituted with 0 to 2 substituents selected fromthe group consisting of halogen, C₁-C₈ alkyl, C₁-C₈ alkoxy, cyano,nitro, hydroxyl, amino and tetrazole) or a pharmaceutically acceptablesalt thereof.
 7. The method according to claim 6, wherein said VLA-4adhesion molecule is formed in connection with an allergic disease. 8.The method of claim 6, wherein the spiro derivative or pharmaceuticallyacceptable salt thereof is a compound, wherein F represents —C(O)—; Arepresents —CH—, carbon having a double bond or nitrogen atom, Brepresents —CH₂— or —NH—; C′ and D represent hydrogen or C′ and Dcooperatively represent O; X and Y independently represent hydrogen,halogen, methyl, methoxy, cyano, nitro, hydroxyl, amino or tetrazole; R₁and R₂ independently represent hydrogen or C₁-C₆ linear alkyl; R₃ and R₄independently represent hydrogen, C₁-C₆ linear alkyl or C₃-C₈ branchedalkyl, or phenyl or benzyl, this phenyl or benzyl being substituted with0 to 2 substituents selected from the group consisting of halogen,methyl, methoxy, cyano, nitro, hydroxyl, amino and tetrazole; when A isa nitrogen atom, R₃, A and R₄ may cooperatively represent (i) saidFormula II (wherein p represents an integer of 0 to 4; E representscarbon atom or nitrogen atom; R₆ and R₇ independently representhydrogen, C₁-C₆ linear alkyl, C₃-C₈ branched alkyl, C₁-C₆ linear acyl,C₃-C₈ branched acyl, pyrrolidinecarbonyl, piperidinecarbonyl, or phenyl,phenylsulfonyl, benzoyl, benzyl, indole or N-phenylamide, this phenyl,phenylsulfonyl, benzoyl, benzyl, indole or N-phenylamide beingsubstituted with 0 to 2 substituents selected from the group consistingof halogen, methyl, methoxy, cyano, nitro, hydroxyl, amino andtetrazole, or said Formula III (wherein R₈ represents hydrogen, C₁C₆linear alkyl, C₃-C₈ branched alkyl, or phenyl or benzyl, this phenyl orbenzyl being substituted with 0 to 2 substituents selected from thegroup consisting of halogen, methyl, methoxy, cyano, nitro, hydroxyl,amino and tetrazole), (ii) said Formula IV (wherein R₉ represents C₁-C₆linear alkyl, C₃-C₈ branched alkyl, C₁-C₆ linear acyl, C₃-C₈ branchedacyl, C₅-C₇ cycloalkylcarbonyl, C₁-C₆ linear alkylsulfonyl, C₃-C₈branched alkylsulfonyl, or benzoyl, phenylsulfonyl or benzyl, thisbenzoyl, phenylsulfonyl or benzyl being substituted with 0 to 2substituents selected from the group consisting of halogen, methyl,methoxy, cyano, nitro, hydroxyl, amino and tetrazole), (iii) saidFormula V: (wherein R₁₀ represents hydrogen, C₁-C₆ linear alkyl, C₃-C₈branched alkyl, C₆-C₁₀ alkylcycloalkyl, or phenyl or benzyl, this phenylor benzyl being substituted with 0 to 2 substituents selected from thegroup consisting of halogen, methyl, methoxy, cyano, nitro, hydroxyl,amino and tetrazole); when A is —CH— or carbon having a double bond, R₃,A and R₄ may cooperatively represent adamantyl or said Formula VI(wherein X₂ and Y₂ represent the same definitions as described above);R₅ represents hydrogen, C₁-C₆ linear alkyl, C₃-C₈ branched alkyl, allyl,homoallyl, C₆-C₁₀ alkylcycloalkyl, or phenyl, benzyl, phenethyl, styrylor naphthylmethyl, this phenyl, benzyl, phenethyl, styryl ornaphthylmethyl being substituted with 0 to 2 substituents selected fromthe group consisting of halogen, methyl, methoxy, cyano, nitro,hydroxyl, amino and tetrazole.
 9. The method of claim 6, wherein thespiro derivative or pharmaceutically acceptable salt thereof is acompound, wherein A represents a nitrogen atom, R₃, A and R₄cooperatively represent (i) said Formula II (wherein R₆ and R₇independently represent hydrogen, C₁-C₃ linear alkyl, C₃-C₆ branchedalkyl, C₁-C₃ linear acyl, C₃-C₆ branched acyl, pyrrolidinecarbonyl,piperidinecarbonyl, or phenyl, benzoyl, benzyl, indole or N-phenylamide,this phenyl, benzoyl, benzyl, indole or N-phenylamide being substitutedwith ₀ to ₂ substituents selected from the group consisting of halogen,methyl, methoxy, cyano, nitro, hydroxyl, amino and tetrazole), or saidFormula III (wherein R₈ represents hydrogen, C₁-C₃ linear alkyl, C₃C₆branched alkyl, or phenyl or benzyl, this phenyl or benzyl beingsubstituted with 0 to 2 substituents selected from the group consistingof halogen, methyl, methoxy, cyano, nitro, hydroxyl, amino andtetrazole), (ii) said Formula IV (wherein R₉ represents C₁-C₃ linearalkyl, C₃-C₆ branched alkyl, C₁-C₃ linear acyl, C₃-C₆ branched acyl,C₅-C₇ cycloalkylcarbonyl, C₁-C₃ linear alkylsulfonyl, C₃-C₈ branchedalkylsulfonyl, or benzoyl or benzyl, this benzoyl or benzyl beingsubstituted with 0 to 2 substituents selected from the group consistingof halogen, methyl, methoxy, cyano, nitro, hydroxyl, amino andtetrazole), or (iii) said Formula V (wherein R₁₀ represents hydrogen,C₁-C₃ linear alkyl, C₃-C₆ branched alkyl, C₆C₁₀ alkylcycloalkyl, orphenyl or benzyl, this phenyl or benzyl being substituted with 0 to 2substituents selected from the group consisting of halogen, methyl,methoxy, cyano, nitro, hydroxyl, amino and tetrazole), the definitionsof the symbols other than mentioned above are the same as thosedescribed in claim 1 or
 2. 10. The method of claim 6, wherein the spiroderivative or pharmaceutically acceptable salt thereof is a compound,wherein A represents a nitrogen atom, R₃, A and R₄ cooperativelyrepresent (i) said Formula II (wherein R₆ and R₇ independently representhydrogen, C₁-C₃ linear acyl, C₃-C₆ branched acyl, pyrrolidinecarbonyl,piperidinecarbonyl, or phenyl, benzoyl, benzyl, indole or N-phenylamide,this phenyl, benzoyl, benzyl, indole or N-phenylamide being substitutedwith 0 to 2 substituents selected from the group consisting of halogen,methyl, methoxy, cyano, nitro, hydroxyl, amino and tetrazole), or saidFormula III (wherein R₈ represents hydrogen, C₁-C₃ linear alkyl, C₃-C₆branched alkyl, or benzyl substituted with 0 to 2 substituents selectedfrom the group consisting of halogen, methyl, methoxy, cyano, nitro,hydroxyl, amino and tetrazole), (ii) said Formula IV (wherein R₉represents C₁-C₃ linear acyl, C₃-C₆ branched acyl, C₅-C₇cycloalkylcarbonyl, or benzoyl or benzyl, this benzoyl or benzyl beingsubstituted with 0 to 2 substituents selected from the group consistingof halogen, methyl, methoxy, cyano, nitro, hydroxyl, amino andtetrazole), or (iii) said Formula V (wherein R₁₀ represents hydrogen,C₁-C₃ linear alkyl, C₃-C₆ branched alkyl, C₆-C₁₀ alkylcycloalkyl, orphenyl or benzyl, this phenyl or benzyl being substituted with 0 to 2substituents selected from the group consisting of halogen, methyl,methoxy, cyano, nitro, hydroxyl, amino and tetrazole), the definitionsof the symbols other than mentioned above are the same as thosedescribed in claim 1 or
 2. 11. A pharmaceutical comprising said spiroderivative or a pharmaceutically acceptable salt thereof according toclaim 3 as an effective ingredient and a carrier.
 12. A pharmaceuticalcomprising said spiro derivative or a pharmaceutically acceptable saltthereof according to claim 4 as an effective ingredient and a carrier.